No full-text available
To read the full-text of this research,
you can request a copy directly from the author.
Glial-neuronal interactions in the mammalian brain
Abstract
Recognition of the importance of glial cells in nervous system functioning is increasing, specifically regarding the modulation of neural activity. This brief review focuses on some of the morphological and functional interactions that take place between astroglia and neurons. Astrocyte-neuron interactions are of special interest because this glia cell type has intimate and dynamic associations with all parts of neurons, i.e., somata, dendrites, axons, and terminals. Activation of certain receptors on astrocytes produces morphological changes that result in new contacts between neurons, along with physiological and functional changes brought about by the new contacts. In response to activation of other receptors or changes in the extracellular microenvironment, astrocytes release neuroactive substances that directly excite or inhibit nearby neurons and may modulate synaptic transmission. Although some of these glial-neuronal interactions have been known for many years, others have been quite recently revealed, but together they are forming a compelling story of how these two major cell types in the brain carry out the complex tasks that mammalian nervous systems perform.
... In mammals, the periodic, synchronous bursts of action potentials in OT neurons drive pulsatile OT release from axon terminals in the neurohypophysis during these periods (Poulain and Wakerley 1982). A range of studies has shown that OT neurons and associated neuroglia undergo dramatic changes during late pregnancy and lactation that could contribute to this specialized activity: changes in morphology, synaptic efficacy, and intrinsic membrane properties (Armstrong 2015;Hatton 2002;Theodosis et al. 2008). ...
... The majority of our results suggested that changes in the sensitivity to TBB are most pronounced in late pregnancy. This corresponds with previous data on synaptic and neuroglial plasticity in the SON (see Armstrong 2015;Hatton 2002;Theodosis et al. 2008, for reviews) as well as our recent findings of a pronounced upregulation of ERK1/2 in OT neurons at late pregnancy (Chandaka et al. 2016). We thus compared blots from SON lysates from virgin and late (E20 -21) pregnant rats for three PP2A subunits (A, B and C), two CK2 subunits (CK2␣ and CK2), and SK3. ...
... We used spike trains that produced different sized I mAHP s to test whether changes in CK2 and PP2A differentially alter I mAHP s in virgins compared with pregnant rats. We examined two stages of pregnancy, E18 -19 and E20 -21, in anticipation that changes in OT reproductive plasticity might occur most significantly later in pregnancy, 24 -48 h before birth (Armstrong 2015;Hatton 2002;Theodosis et al. 2008). ...
Oxytocin (OT) neurons exhibit larger afterhyperpolarizations (AHPs) following spike trains during pregnancy and lactation, when these neurons burst and release more OT. Calcium-dependent AHPs mediated by SK channels show this plasticity, and are reduced when the channel is phosphorylated by casein kinase 2 (CK2), and increased when dephosphorylated by protein phosphatase 2A (PP2A), by altering Ca2+sensitivity. We compared AHP currents in supraoptic OT neurons after CK2 inhibition with 4,5,6,7-tetrabromobenzotriazole (TBB), or PP1-PP2A inhibition with okadaic acid (OA) focusing on the peak current at 100 ms representing the SK-mediated, medium AHP (ImAHP). In slices from virgin and two groups of pregnant rats (E18-19, or E20-21), ImAHPswere evoked with 3, 10, and 17 spike trains (20 Hz). With 3-spike trains, TBB increased the ImAHPthe greatest extent in virgin compared to both groups of pregnant animals. A difference between virgins and E20-21 rats was evident with a 10-spike train but the increases in ImAHPswere similar among groups with 17-spike trains. In contrast, OA consistently reduced the ImAHPin all cases, but showed no differential effects among groups. In Western blots, CK2, CK2, PP2A-A, PP2A-B and PP2A-C were found in supraoptic lysates, and expression of CK2α and CK2β were reduced in E20-21 rats. Co-immunoprecipitation revealed that calmodulin, CK2α, and PP2A-C were associated with SK3 protein. The results suggest that a downregulation of SK3-associated CK2 during late pregnancy increases the sensitivity of the SK calmodulin (Ca2+) sensor for ImAHP, contributing to the enhanced ImAHP.
... Thus astrocytes represent anatomically an obligatory interface between neurons and extraparenchymal cells. functions (Althaus and Landsberg, 2000), undergo morphological changes ultimately leading to neuroendocrine regulation in the hypothalamus (Hatton, 2002), intervene in maintenance and formation of synapses (Ullian et al., 2001), influence the blood brain barrier properties (Rubin and Staddon, 1999), participate in the immune response and inflammation, produce chemokines attracting inflammatory cells, are involved in apoptosis and phagocytosis of inflammatory cells, influence the differentiation of immune cells and the proliferation of most brain cell types, and may serve as NSC to replace glia and neurons throughout adulthood . ...
... The anatomical organization of the hypothalamic portion of the mHNS is schematically illustrated in figure 11A (coronal view). Astrocytes display intimate contacts with the magnocellular neurons at every level of the neuronal organization (see structure in Mercier and Hatton, (2000) and ultrastructure in Hatton, (2002). The most impressive anatomical portion of the mHNS organization illustrating meningeal-neuronal-astrocytic interactions is the supraoptic nucleus. ...
... When physiological conditions such as dehydration, lactation arouse the mHNS, morphological and functional changes occur in the supraoptic nucleus (Hatton, 2002). The plastic changes in the core and dendritic zone (more ventral portion) of the supraoptic nucleus consist of astrocytic retraction. ...
The current view is that glial cells play important roles in brain functions. Previous authors have shown that glial cells (primarily astrocytes) are connected by gap junctions and functionally cooperate to form a network and coordinate interactions with neurons (see Chapter 13). On the basis of newly revealed anatomical data (Mercier and Hatton, 2000; 2001, Mercier et al., 2002; 2003) and re-analysis of the literature, it is here proposed that the glial network is an anatomical and functional interface between the neuronal compartment and meninges. Meninges are widely thought to be merely protective membranes that do not serve any important physiologically relevant function during adulthood. However, meninges produce numerous cytokines and growth factors (GF) throughout adulthood. Numerous of these signaling molecules bind to heparan sulfate proteoglycans (HSPG) and/or interact with other extracellular matrix proteins contained in the basal lamina (BL) that is systematically interposed between meningeal cells and astrocytes of the glia limitans (GL). Therefore, meningeal-produced cytokines and GF can directly influence glial cell functions. Similar mechanisms have been demonstrated during development between the connective tissues and parenchymal tissues or epithelia (mesodermo-neuroectodermal or mesodermo-epithelial induction). Astrocytes produce cytokines and GF, likely in response to the information they receive from meningeal and vascular cells. This may form the basis of signaling cascades from the meninges to neurons via astrocytes.
... Astrocytes are known to be important components in the regulation of AVP release 48 . Hyperosmotic challenges retract the astrocytic processes surrounding AVP neurons, which facilitates neuronal interactions through synapses and dendritic gap junctions, increasing AVP neuronal excitability and promoting synchronous AVP secretion 49 . In contrast, hypoosmotic challenges inhibit AVP neuronal activity by evoking the expansion of astrocytic processes, wedging them in between neuronal dendrites and lessening dendritic gap junctional connectivity 49 . ...
... Hyperosmotic challenges retract the astrocytic processes surrounding AVP neurons, which facilitates neuronal interactions through synapses and dendritic gap junctions, increasing AVP neuronal excitability and promoting synchronous AVP secretion 49 . In contrast, hypoosmotic challenges inhibit AVP neuronal activity by evoking the expansion of astrocytic processes, wedging them in between neuronal dendrites and lessening dendritic gap junctional connectivity 49 . In the present study, AVP neuron-specific BiP knockdown led not only to the death of AVP neurons but also to the activation of astrocytes. ...
The immunoglobulin heavy chain binding protein (BiP), also referred to as 78-kDa glucose-regulated protein (GRP78), is a pivotal endoplasmic reticulum (ER) chaperone which modulates the unfolded protein response under ER stress. Our previous studies showed that BiP is expressed in arginine vasopressin (AVP) neurons under non-stress conditions and that BiP expression is upregulated in proportion to the increased AVP expression under dehydration. To clarify the role of BiP in AVP neurons, we used a viral approach in combination with shRNA interference for BiP knockdown in mouse AVP neurons. Injection of a recombinant adeno-associated virus equipped with a mouse AVP promoter and BiP shRNA cassette provided specific BiP knockdown in AVP neurons of the supraoptic (SON) and paraventricular nuclei (PVN) in mice. AVP neuron-specific BiP knockdown led to ER stress and AVP neuronal loss in the SON and PVN, resulting in increased urine volume due to lack of AVP secretion. Immunoelectron microscopy of AVP neurons revealed that autophagy was activated through the process of AVP neuronal loss, whereas no obvious features characteristic of apoptosis were observed. Pharmacological inhibition of autophagy by chloroquine exacerbated the AVP neuronal loss due to BiP knockdown, indicating a protective role of autophagy in AVP neurons under ER stress. In summary, our results demonstrate that BiP is essential for the AVP neuron system.
... Using comparative ultrastructural analyses, it was discovered that direct appositions among HNS neurons normally separated by astroglial processes vary considerably under different conditions of neurosecretion Hatton, 1976, 1977;Theodosis et al., 1981;Theodosis and Poulain, 1984;Hatton et al., 1984). While most of the work has been done on OT neurons, and particularly during the reproductive cycle (reviewed in Hatton, 2002;Theodosis et al., 2008), neuroglial interactions have also been reported among VP neurons during dehydration (Gregory et al., 1980;Marzban et al., 1992;Miyata et al., 1994), although this was not found in all studies (see Chapman et al., 1986). ...
... Although some have suggested an increase in gap junctional coupling between magnocellular neurons during enhanced hormone release, this is based on increased amounts of dye coupling (see Hatton, 2002 for review) and the increased presence of connexin 32 (Micevych et al., 1996). As stated before, extensive ultrastructural analyses support that gap junctions are probably restricted to astrocytes, and there is little evidence for electrotonic coupling in basal conditions (Yang and Hatton, 1988). ...
Oxytocin (OT) and vasopressin (VP) are synthesized principally by magnocellular neuroendocrine cells in the hypothalamic supraoptic and paraventricular nuclei and are secreted systemically by axonal projections to the neurohypophysis. Blood-borne OT and VP act peripherally to control reproductive function and fluid homeostasis. OT and VP also act centrally to regulate behavior, as well as locally via dendritic release to regulate OT/VP neuron activity via autocrine/paracrine actions. The anatomical organization of the magnocellular neuroendocrine systems is highly plastic and undergoes dramatic state-dependent alterations in physiology and hormone secretion. OT and VP neuroendocrine cells are excellent models of physiological state-dependent regulation of neurosecretion.
... Additionally, parvocellular OT neurons send their axons not only to the neurohypophysis, but also to a variety of central sites (Gimpl and Fahrenholz 2001). Furthermore, based on IHC (immunohistochemistry), ISHH (in situ hybridization histochemistry) and Arrays (DNA microarrays) detection, co-localization of OT and HA (H1 and H3) receptors occurs in the OT neurons in the PVN and SON (Georgina et al. 2012;Hatton 2002;Lemonick et al. 2001;Markovic and Challiss 2009). In addition, arrays detection has shown that co-location of OT and β2adrenergic receptors takes place in the OT neurons in the SON (Georgina et al. 2012;Hatton 2002). ...
... Furthermore, based on IHC (immunohistochemistry), ISHH (in situ hybridization histochemistry) and Arrays (DNA microarrays) detection, co-localization of OT and HA (H1 and H3) receptors occurs in the OT neurons in the PVN and SON (Georgina et al. 2012;Hatton 2002;Lemonick et al. 2001;Markovic and Challiss 2009). In addition, arrays detection has shown that co-location of OT and β2adrenergic receptors takes place in the OT neurons in the SON (Georgina et al. 2012;Hatton 2002). So, it is more likely that the interaction of histaminergic and noradrenergic systems with oxytocin on feeding behavior in broilers occurs in SON and PVN. ...
Oxytocin neurons have a physiological role in food intake and energy balance. Several studies have shown that central histaminergic and adrenergic systems synapse on oxytocin neurons but there is no information for their interaction on food intake regulation in birds. The purpose of this study was to examine the effects of intracerebroventricular (ICV) injection of α-fluoromethylhistidine (α-FMH, histidine decarboxylase inhibitor), chlorpheniramine (histamine H1 receptors antagonist), famotidine (histamine H2 receptors antagonist), thioperamide (histamine H3 receptors antagonist), prazosin (α1 receptor antagonist), yohimbine (α2 receptor antagonist), metoprolol (β1 adrenergic receptor antagonist), ICI 118,551 (β2 adrenergic receptor antagonist) and SR59230R (β3 adrenergic receptor antagonist) on oxytocin-induced hypophagia in 3-h food-deprived (FD3) neonatal broiler chicken. In Experiment 1, 3 h-fasted chicks were given an ICV injection of saline, α-FMH (250 nmol), oxytocin (10 μg) and co-injection of α-FMH + oxytocin. Experiments 2-9 were similar to experiment 1 except birds were injected with chlorpheniramine (300 nmol), famotidine (82 nmol), thioperamide (300 nmol), prazosin (10 nmol), yohimbine (13 nmol), metoprolol (24 nmol), ICI 118,551(5 nmol) and SR59230R (20 nmol) instead of α-FMH, respectively. After injection cumulative food intake was measured until 120 min post injection. According to the results, ICV injection of oxytocin significantly decreased food intake in broiler chickens (P < 0.001). ICV injection of α-FMH significantly attenuated hypophagic effect of oxytocin (P < 0.001). Also, co-injection of chlorpheniramine plus oxytocin significantly decreased the effect of oxytocin on food intake (P < 0.001). Co-administration of thioperamide and oxytocin significantly amplified hypophagic effect of oxytocin in chickens (P < 0.001). In addition, ICI 118,551 attenuated hypophagic effect of oxytocin (P < 0.001); while famotidine, prazosin, yohimbine, metoprolol and SR59230R had no effect on oxytocin- induced food intake in FD3 broiler chickens. These results suggest that the effect of oxytocin on food intake is probably mediated by histaminergic (via H1 and H3 receptors) and noradrenergic (via β2 receptors) systems in broiler chickens.
... Estudis recents han definit que un sol astròcits està en contacte amb 4-8 somes neuronals i amb 300-600 dendrites procedents de diferents neurones (figura 5) (Benarroch, 2005). Figura 5. Micrografia electrònica on es mostra un astròcit (blau) en contacte amb terminals axonals (At) fent sinapsis asimètriques amb 4 espines (sp1, sp2, i dos no marcades) (Hatton, 2002). ...
... Els astròcits secreten molècules que promouen o inhibeixen el creixement axonal, proporcionant una guia pel desenvolupament neuronal. També intervenen en el reemplaçament neuronal a partir de l'estoc de cèl·lules mare neurals (Blackburn et al., 2009;Hatton, 2002;Kanemaru et al., 2007). ...
... Given the importance of glial-neuronal interactions for normal CNS functioning (Hatton, 2002), we hypothesized that established co-cultures of neurons and glia may be less susceptible to the toxic effects by chronic ethanol exposure and more specifically, the toxicity observed during ethanol withdrawal in that we would see little to no astrocytic or neuronal cell death. The purpose of the current study was to assess the effect of continuous ethanol exposure (CEE) on cell viability of co-cultured astrocytes and cerebellar granule cells (CGCs), as well as changes in NMDA-mediated currents (I NMDA ), during ethanol intoxication and throughout the ethanol withdrawal phase. ...
... A large percentage of cells contained in the CNS are astrocytes, and many normal as well as abnormal brain functions are due to interactions among glia and neurons (Hatton, 2002). Yet, to date, in vitro chronic ethanol studies have assessed the effects of ethanol only on pure astrocytic or neuronal cultures and in many instances, ethanol treatment is begun immediately after plating (Pascual et al., 2003;Gausch et al., 2003). ...
Long-term ethanol exposure has deleterious effects on both glial and neuronal function. We assessed alterations in both astrocytic and neuronal viability, and alterations in N-methyl-d-aspartate receptor (NMDAR) function, in cocultures of rat cerebellar granule cells (CGCs) and astrocytes after continuous ethanol exposure (CEE). Treatment of cells with 100 mM EtOH once every 24 h for 4 days resulted in a mean ethanol concentration of 57.3 ± 2.1 mM. Comparisons between control and post-ethanol-treated cells were made 4 days after the last ethanol treatment. CEE did not alter glial cell viability, as indicated by the absence of either changes in astrocytic morphology, actin depolymerization, or disruption of astrocytic intracellular mitochondrial distribution at any day postethanol treatment. The CGCs were healthy and viable after CEE, as indicated by phase-contrast microscopy and the trypan-blue exclusion method. Whole-cell patch-clamp experiments indicated that NMDA-induced currents (I(NMDA)) were altered by CEE treatment. Similar to previous results obtained during the withdrawal phase from chronic ethanol exposure, I(NMDA) from CEE-treated cells were significantly larger than I(NMDA) from NMDARs in control CGCs, but returned to control values by the fourth day post-CEE. However, after the last ethanol dosing and during a time when ethanol concentrations remained high, I(NMDA) were significantly smaller than control values. Identical results were observed in CGCs expressing the NR2A or NR2B subunit. In summary, both neurons and astrocytes remained healthy following exposure to CEE with no signs of neurotoxicity at the cellular level, and modulation of NMDAR function is consistent with findings from prior experiments. Thus, we conclude that the CEE paradigm in glial-neuronal cocultures readily lends itself to long-term in vitro studies of ethanol effects that include glial-neuronal interactions and the ability to study ethanol withdrawal-induced neurotoxicity.
... In the stratum radiatum of hippocampal CA1 region, 57% of the synapses formed between Schaffer collaterals and CA1 pyramidal neurones are in contact with astrocytes [23] . However, the degree of synaptic coverage by glia is a dynamic process, as best described in the magnocellular hypothalamo-neurohypophysial system [24] . Neurone-glia signalling can also occur through direct synapse-like connections synaptoids, ectopic sites at pre-synaptic terminals or even bona fide synaptic contacts ( Figure 2C-2E). ...
... Note postsynaptic densities in glial elements. Scale bars, 100 nm in B (50 nm inset), 330 nm in C, 100 nm in D, and 250 nm in E. Micrograph in A is modified from [111] and reprinted from [24] , with permission from American Physiological Society. Micrograph in B is adapted by permission from Macmillan Publishers Ltd: Nature Neuroscience [112] , copyright 2004. ...
Our view of astrocytes in the operation of the brain is changing dramatically over the last 3 decades. Astroglial calcium excitability controls the release of gliotransmitters, which can occur at the tripartite synapse. Astrocytes not only modulate synaptic transmission by releasing and taking up transmitters, but also receiving neuronal signals that act upon astrocytic plasma membrane receptors. This process represents the bidirectional neurone-glia communication. Additionally, astrocytes play role in the regulation of blood flow as well as ion and water homeostasis. Many of the brain dysfunctions are primary astropathies, including hepatic encephalopathy and Alexander disease, while other brain malfunctions, such as epilepsy and Alzheimer disease, may have substantial astrocytic contribution. Thus, these star-shaped cells by their roles in (patho)physiology of the brain seem to live up to the expectation one can have from their given name - astrocyte.
... Thus, VRAC, LRRC8A, and taurine, seem to play a critical role in astrocytic homeostasis. Moreover, previous studies have shown that the highest levels of taurine in the brain are found in cerebral cortex, hippocampus, caudate-putamen, cerebellum, and in hypothalamic supraoptic nucleus (Hussy et al., 1997(Hussy et al., , 2000Hatton, 2002). VRAC and VRAC-like currents have been reported in mice and rats in several brain regions including cervical sympathetic ganglions, CA1 region of the hippocampus, cerebral cortex, hypothalamus, and cerebellum (Leaney et al., 1997;Patel et al., 1998;Inoue et al., 2005;Sato et al., 2011;Zhang et al., 2011). ...
... Thus, VRAC, LRRC8A, and taurine, seem to play a critical role in astrocytic homeostasis. Moreover, previous studies have shown that the highest levels of taurine in the brain are found in cerebral cortex, hippocampus, caudate-putamen, cerebellum, and in hypothalamic supraoptic nucleus (Hussy et al., 1997(Hussy et al., , 2000Hatton, 2002). VRAC and VRAC-like currents have been reported in mice and rats in several brain regions including cervical sympathetic ganglions, CA1 region of the hippocampus, cerebral cortex, hypothalamus, and cerebellum (Leaney et al., 1997;Patel et al., 1998;Inoue et al., 2005;Sato et al., 2011;Zhang et al., 2011). ...
Taurine is considered the most abundant free amino acid in the brain. Even though there are endogenous mechanisms for taurine production in neural cells, an exogenous supply of taurine is required to meet physiological needs. Taurine is required for optimal postnatal brain development; however, its brain concentration decreases with age. Synthesis of taurine in the central nervous system (CNS) occurs predominantly in astrocytes. A metabolic coupling between astrocytes and neurons has been reported, in which astrocytes provide neurons with hypotaurine as a substrate for taurine production. Taurine has antioxidative, osmoregulatory, and anti-inflammatory functions, among other cytoprotective properties. Astrocytes release taurine as a gliotransmitter, promoting both extracellular and intracellular effects in neurons. The extracellular effects include binding to neuronal GABAA and glycine receptors, with subsequent cellular hyperpolarization, and attenuation of N-methyl-D-aspartic acid (NMDA)-mediated glutamate excitotoxicity. Taurine intracellular effects are directed toward calcium homeostatic pathway, reducing calcium overload and thus preventing excitotoxicity, mitochondrial stress, and apoptosis. However, several physiological aspects of taurine remain unclear, such as the existence or not of a specific taurine receptor. Therefore, further research is needed not only in astrocytes and neurons, but also in other glial cells in order to fully comprehend taurine metabolism and function in the brain. Nonetheless, astrocyte’s role in taurine-induced neuroprotective functions should be considered as a promising therapeutic target of several neuroinflammatory, neurodegenerative and psychiatric diseases in the near future. This review provides an overview of the significant relationship between taurine and astrocytes, as well as its homeostatic and neuroprotective role in the nervous system.
... Astrocytes are the largest population of cells in the mammalian brain, where they provide lifelong homeostatic support to neurons [8]. Interestingly, astrogliosis, an inflammatory response seen after brain injury and in some neurodegenerative diseases, involves the activation and proliferation of astrocytes and their eventual occupation of injured brain areas [9][10][11]. ...
Astrocyte-to-neuron reprogramming is a promising therapeutic approach for treatment of neurodegenerative diseases. The use of small molecules as an alternative to the virus-mediated ectopic expression of lineage-specific transcription factors negates the tumorigenic risk associated with viral genetic manipulation and uncontrolled differentiation of stem cells. However, because previously developed methods for small-molecule reprogramming of astrocytes to neurons are multistep, complex, and lengthy, their applications in biomedicine, including clinical treatment, are limited. Therefore, our objective in this study was to develop a novel chemical-based approach to the cellular reprogramming of astrocytes into neurons with high efficiency and low complexity. To accomplish that, we used C8-D1a, a mouse astrocyte cell line, to assess the role of small molecules in reprogramming protocols that otherwise suffer from inconsistencies caused by variations in donor of the primary cell. We developed a new protocol by which a chemical mixture formulated with Y26732, DAPT, RepSox, CHIR99021, ruxolitinib, and SAG rapidly and efficiently induced the neural reprogramming of astrocytes in four days, with a conversion efficiency of 82 ± 6%. Upon exposure to the maturation medium, those reprogrammed cells acquired a glutaminergic phenotype over the next eleven days. We also demonstrated the neuronal functionality of the induced cells by confirming KCL-induced calcium flux.
... Astrocytic processes form parts of the blood brain barrier [4,25] and regulate the uptake and distribution of metal ions within the brain [24,41,67,89,90], which makes them the first parenchymal cell type that will get in contact with iron after having crossed the blood brain barrier [41]. Due to their large intracellular iron pool together with their close contacts to various other brain cell types [3,35,39,67,89], they play a key role in processes of iron distribution within the neuropil [68]. The cellular iron concentration in astrocytes remains unchanged in PD, which may indicate that they are largely able to maintain their homeostatic pathways, even under inflammatory conditions. ...
Iron is essential for neurons and glial cells, playing key roles in neurotransmitter synthesis, energy production and myelination. In contrast, high concentrations of free iron can be detrimental and contribute to neurodegeneration, through promotion of oxidative stress. Particularly in Parkinson’s disease (PD) changes in iron concentrations in the substantia nigra (SN) was suggested to play a key role in degeneration of dopaminergic neurons in nigrosome 1. However, the cellular iron pathways and the mechanisms of the pathogenic role of iron in PD are not well understood, mainly due to the lack of quantitative analytical techniques for iron quantification with subcellular resolution. Here, we quantified cellular iron concentrations and subcellular iron distributions in dopaminergic neurons and different types of glial cells in the SN both in brains of PD patients and in non-neurodegenerative control brains (Co). To this end, we combined spatially resolved quantitative element mapping using m icro p article i nduced X -ray e mission (µPIXE) with nickel-enhanced immunocytochemical detection of cell type-specific antigens allowing to allocate element-related signals to specific cell types. Distinct patterns of iron accumulation were observed across different cell populations. In the control (Co) SNc, oligodendroglial and astroglial cells hold the highest cellular iron concentration whereas in PD, the iron concentration was increased in most cell types in the substantia nigra except for astroglial cells and ferritin-positive oligodendroglial cells. While iron levels in astroglial cells remain unchanged, ferritin in oligodendroglial cells seems to be depleted by almost half in PD. The highest cellular iron levels in neurons were located in the cytoplasm, which might increase the source of non-chelated Fe ³⁺ , implicating a critical increase in the labile iron pool. Indeed, neuromelanin is characterised by a significantly higher loading of iron including most probable the occupancy of low-affinity iron binding sites. Quantitative trace element analysis is essential to characterise iron in oxidative processes in PD. The quantification of iron provides deeper insights into changes of cellular iron levels in PD and may contribute to the research in iron-chelating disease-modifying drugs.
... Abbreviations: 3rd V the third cerebral ventricle, A. Pit anterior pituitary, ME median eminence, mPVN magnocellular division of the PVN, OC optic chiasm, Pit pituitary, P. Pit posterior pituitary, pPVN parvocellular division of the PVN, SON supraoptic nucleus Among many extracellular factors, astrocytic plasticity or morphological and functional change plays a pivotal role since many external regulatory factors are under intensive modulation of astrocytes adjacent to MNCs. This astrocytic plasticity has been identified in studies on dehydration and lactation since half a century ago [2,3]. Recently, knowledge of astrocytic-MNC interactions increases significantly with progress of in vitro studies on osmotic challenges and lactation with modern technology. ...
Studies on the interactions between astrocytes and neurons in the hypothalamo-neurohypophysial system have significantly facilitated our understanding of the regulation of neural activities. This has been exemplified in the interactions between astrocytes and magnocellular neuroendocrine cells (MNCs) in the supraoptic nucleus (SON), specifically during osmotic stimulation and lactation. In response to changes in neurochemical environment in the SON, astrocytic morphology and functions change significantly, which further modulates MNC activity and the secretion of vasopressin and oxytocin. In osmotic regulation, short-term dehydration or water overload causes transient retraction or expansion of astrocytic processes, which increases or decreases the activity of SON neurons, respectively. Prolonged osmotic stimulation causes adaptive change in astrocytic plasticity in the SON, which allows osmosensory neurons to reserve osmosensitivity at new levels. During lactation, changes in neurochemical environment cause retraction of astrocytic processes around oxytocin neurons, which increases MNC’s ability to secrete oxytocin. During suckling by a baby/pup, astrocytic processes in the mother/dams exhibit alternative retraction and expansion around oxytocin neurons, which mirrors intermittently synchronized activation of oxytocin neurons and the post-excitation inhibition, respectively. The morphological and functional plasticities of astrocytes depend on a series of cellular events involving glial fibrillary acidic protein, aquaporin 4, volume regulated anion channels, transporters and other astrocytic functional molecules. This review further explores mechanisms underlying astroglial regulation of the neuroendocrine neuronal activities in acute processes based on the knowledge from studies on the SON.
... Organisation du système hypothalamoneurohypophysaire d'après(Hatton 2002) afférences axoniques(Armstrong et al 1982). Deux populations neuronales distinctes composent ces noyaux : l'une exprime l'ocytocine (OT) et l'autre la vasopressine (AVP). ...
... After 24 hrs. of dehydration, L-channel Ca 2+ density accounts for an increase in Ca 2+ current density in both OT and VP neurons (125). Water deprivation or salt loading not only activates both OT and VP neurons, but also leads to many morphological, electrophysiological and synaptic changes in magnocellular neurons and associated glia similar to those observed specifically in OT neurons during lactation (126)(127)(128). ...
To understand the contribution of intrinsic membrane properties to the different firing patterns of oxytocin (OT) and vasopressin (VP) neurons in vivo, in vitro studies are needed, where stable intracellular recordings can be made. Combining immunochemistry for OT and VP and intracellular dye injections allows characterization of identified OT and VP neurons, and several differences between the two cell types have emerged. These include a greater transient K⁺ current that delays spiking to stimulus onset, and a higher Na⁺ current density leading to greater spike amplitude and a more stable spike threshold, in VP neurons. VP neurons also show a greater incidence of both fast and slow Ca²⁺‐dependent depolarizing afterpotentials, the latter of which summate to plateau potentials and contribute to phasic bursting. In contrast, OT neurons exhibit a sustained outwardly rectifying potential (SOR), and a consequent depolarizing rebound potential, not found in VP neurons. The SOR makes OT neurons more susceptible to spontaneous inhibitory synaptic inputs and correlates with a longer period of spike frequency adaptation in these neurons. While both types exhibit prominent Ca²⁺‐dependent afterhyperpolarizing potentials (AHPs) that limit firing rate and contribute to bursting patterns, Ca²⁺‐dependent AHPs in OT neurons selectively show significant increases during pregnancy and lactation. In OT, but not VP neurons, AHPs are highly dependent on the constitutive presence of the second messenger, phosphatidylinositol 4,5‐bisphosphate, which permissively gates N‐type channels that contribute the Ca²⁺ during spike trains that activates the AHP. In contrast to the intrinsic properties supporting phasic bursting in VP neurons, the synchronous bursting of OT neurons has only been demonstrated in vitro in cultured hypothalamic explants and is completely dependent on synaptic transmission. Additional differences in Ca²⁺ channel expression between the two neurosecretory terminal types suggests these channels are also critical players in the differential release of OT and VP during repetitive spiking, in addition to their importance to the potentials controlling firing patterns.
This article is protected by copyright. All rights reserved.
... There is evidence of β-adrenergic receptor on fibrous and protoplasmic astrocytes present in the cerebral cortex, neostriatum, gray matter and the white matter . Such noradrenergic innervation in astrocytes may be responsible for the regulation of morphological plasticity (Hatton, 2002;Monlezun et al., 2005), energy metabolism (Bolanos and Almeida, 2006), membrane transport (Hennemann et al., 2001), gap junction permeability (Saez et al., 2005) and immunological responses (Dong and Benveniste, 2001) in these cells. Noradrenergic effects on astrocytes are essential during consolidation of episodic, long-term memory, which is reinforced by β-adrenergic activation (Hertz et al., 2004). ...
Thyroid hormones (TH) have profound effects on brain development both in vivo and in vitro. Deficiency of the hormones severely impair the various stages of neuronal development like proliferation, differentiation, axonal migration, myelinogenesis, synaptogenesis etc. Other neural cells like oligodendroglia, astroglia also respond to the hormones as they bear TH receptors. Astrocytes undergo dynamic changes in morphology during normal brain development. Such changes in astrocyte morphology also occur during various pathophysiological changes. Previous workers from this laboratory had established an in vitro long-term primary culture of astrocytes derived from neonatal rat brains covering the entire span of morphological maturation of astrocytes and demonstrated the successive changes in morphology of the cells from radial glia to flat polygonal cells with epithelioid morphology and finally to mature process bearing cells with stellate morphology. It was observed that deficiency of TH delayed the first step and completely prevented the final step of differentiation. In another study it was demonstrated that -adrenergic receptor (-AR) antagonists could completely block TH-induced morphogenesis of astrocytes.
With this background, the present work was conceived. Two major issues were addressed. We investigated the possible signaling mechanisms associated with TH-induced differentiation of astrocytes. We also studied the effect of TH-induced differentiation on the -adrenergic system in astrocytes.
In this dissertation work, it was observed that, TH initially activates protein kinase A (PKA), with a peak activity at 2 hr, which then falls back to basal level. Although there is no visible change in morphology of the cells during this time, this activation of PKA was sufficient to drive the process of transformation to completion. It was further observed that TH treatment resulted in a biphasic response on cellular p-MAP kinase (p-ERK) level, an initial decline in p-ERK level followed by an induction at 18-24 hr, which in turn activate p-CREB. Moreover, the induction in p-ERK level coincides with initiation of morphological differentiation of the astrocytes. Contrary to a short-term decline in p-ERK activity during TH treatment, a long lasting activation of p-ERK activity at a later stage appeared to be critical for the transformation of astrocytes. Ontogenic studies showed a gradual increase in the specific binding of 125I-PIN to β2-AR in response to TH treatment due to the increased affinity of the receptors. Although, β2-AR didn’t shown any significant change in the transcriptional level, overexpression of the same gene in the hypothyroid astrocyte caused morphological transformation even in absence of TH. While -arrestin showed a steady increase in total protein level as well as in mRNA level after TH-treatment, an initial decrease in 125I-PIN binding to β2-AR was also encountered during early phase (2 – 12 hr) of hormone exposure which coincides with a decline in phospho--arrestin protein level at the same time. The result indicates a regulatory influence of the hormone in increasing the affinity of the said receptor and a putative role of its regulator in TH-induced morphological differentiation of astrocytes.
... Interactions between astrocytes and neurons are essential for several brain functions, including homeostatic and metabolic signalling and trophic and pathophysiological regulation (Hatton, 2002;Barker and Ullian, 2010). The hypothalomo-neurohypophyseal system clearly demonstrates the importance of these functional interactions. ...
Among the various functions of the hypothalamo-neurohypophyseal system, it is particularly at stake in the control pathways of plasma osmolality and is especially stimulated during dehydration of the body. This system receives peripheral afferents, such as those from the carotid sinus, but also information intrinsic to the central nervous system (CNS) from circumventricular organs. The control system of osmoregulation gets through these structures and information such as plasma osmolality may be quickly assessed to adjust the level of release of arginine vasopressin (AVP), an anti-diuretic hormone that controls the reabsorption of water in the kidney. The supraoptic nucleus (SON), composed of magnocellular neurons synthesizing AVP, thus receives many afferents that modulate its activity, such as afferents involving noradrenaline (NA), as well as aminoacidergic afferents that release glutamate, aspartate and GABA. Our team is interested in the mechanisms regulating the expression and release of AVP by noradrenergic afferents. The effect of norepinephrine passes through a nitrergic pathway (NO) to control the expression of AVP (Grange-Messent et al, 2004). In addition, the work of S. Parsadaniantz Malik and his team have shown that the expression of chemokines, including chemokine SDF1 and its receptor CXCR4, increased during dehydration (Callewaere et al., 2006). This demonstration of the presence of chemokines in the magnocellular neurons and their involvement in dehydration and the results we obtained on NO suggests that inflammatory mediators are endogenous signalling molecules involved in the signalling chain involved in osmoregulation. The objective of this thesis is to focus on the oxidative metabolism in the magnocellular nuclei, since in the process of post-lesional plasticity, changes in the expression of inflammatory mediators are accompanied by changes in the metabolism and the production of free radicals or reactive oxygen species (ROS). Our study model is the adult C3H/HeJ mice subjected to plasma hyperosmolarity. This paradigm is known to stimulate the osmoregulatory axis and to cause an increase in AVP without causing stress. In the case of a chronic hyperosmolar stimulation, the results show that upon activation of the osmoregulatory axis, demonstrated by the expression of the protein c-Fos, AVP synthesis is accompanied by a production of ROS, taken in charge by superoxide dismutase type 2 (SOD 2) and catalase. Measurements of plasma osmolality indicate that osmoregulatory response exerted by AVP is set up gradually and only after 8 days of osmotic stimulation, the system returns to the control level showing a new allostatic state. In the case of an acute hyperosmolar stimulation, ROS were produced in the early phase of hyperosmolarity demonstrated by the expression of c-Fos, and are taken in charge by SOD 2 and catalase. In addition, the production of ROS is essential for the increase in AVP synthesis in response to plasma hyperosmolarity, since the presence of an antioxidant, alpha-lipoic acid (ALA) administered by osmotic stimulation, there is an inhibition of AVP synthesis. The second part of my thesis seeks to determine the role of ROS in the control exerted by the noradrenergic afferents via the nitric oxide (NO). The results show that transgenic mice TG8, which have high rates in NA in the CNS, have levels of ROS production in the NSO higher than their non-transgenic controls, C3H/HeJ mice, giving an indication that noradrenergic pathway is involved in vivo. To confirm the role of ROS in the noradrenergic signaling pathway, we analyzed ex vivo the effect of NA on hypothalamic slices maintained in survival. This allowed us also to further clarify the relative contributions of NO and ROS in the noradrenergic regulation of the expression of AVP. These results demonstrate for the first time the importance of ROS as endogenous signal molecules involved in osmotic regulation. From this study, a new and original role of ROS as mediators of physiological intracellular signaling pathways emerges.
... Advances in neural cell culture have strengthened biology knowledge, particularly in the structure, morphology and development of different cell types, as well as neuroprotective or neurotoxic effects of various substances; however, the interaction between neuronal and glial cells such as astrocytes, which are critical in the physiological interdependence mediated by chemical signals such as trophic factors, in the formation of synapses, and providing structural support, homeostasis and other functions that support the development, growth and neuronal activity, are rarely taken into account (Fu et al., 2013;Hatton, 2002). ...
... Cell culture Human brain vascular smooth muscle cells (BVSMCs; Schwartz et al., 1986;Braun et al., 1999) and human astrocytes (van der Laan et al., 1997;Hatton, 2002;Chen and Swanson, 2003), obtained from ScienCell Research Laboratory, were cultured in an incubator with 5% CO 2 /95% air at 37°C. The second or third passages of the cells were used for experiments. ...
Astrocytes express laminin and assemble basement membranes (BMs) at their endfeet, which ensheath the cerebrovasculature. The function of astrocytic laminin in cerebrovascular integrity is unknown. We show that ablation of astrocytic laminin by tissue-specific Cre-mediated recombination disrupted endfeet BMs and led to hemorrhage in deep brain regions of adult mice, resembling human hypertensive hemorrhage. The lack of astrocytic laminin led to impaired function of vascular smooth muscle cells (VSMCs), where astrocytes have a closer association with VSMCs in small arterioles, and was associated with hemorrhagic vessels, which exhibited VSMC fragmentation and vascular wall disassembly. Acute disruption of astrocytic laminin in the striatum of adult mice also impaired VSMC function, indicating that laminin is necessary for VSMC maintenance. In vitro, both astrocytes and astrocytic laminin promoted brain VSMC differentiation. These results show that astrocytes regulate VSMCs and vascular integrity in small vessels of deep brain regions. Therefore, astrocytes may be a possible target for hemorrhagic stroke prevention and therapy.
... The cytoskeleton composition of astrocytic processes, especially that of PAPs, is not well-studied and controversially discussed in the literature (Safavi-Abbasi et al., 2001; Hatton, 2002; Potokar et al., 2007; Peng et al., 2008; Kreft et al., 2009; Ou et al., 2009). To our best knowledge, this is the first time that cytoskeletal components are unambiguously localized in the different compartments of in situ like astrocytes. ...
Studying the distribution of astrocytic antigens is particularly hard when they are localized in their fine, peripheral astrocyte processes (PAPs), since these processes often have a diameter comparable to vesicles and small organelles. The most appropriate technique is immunoelectron microscopy, which is, however, a time-consuming procedure. Even in high resolution light microscopy, antigen localization is difficult to detect due to the small dimensions of these processes, and overlay from antigen in surrounding non-glial cells. Yet, PAPs frequently display antigens related to motility and glia-synaptic interaction. Here, we describe the dissociation of morphologically intact glial cells (DIMIGs), permitting unambiguous antigen localization using epifluorescence microscopy. Astrocytes are dissociated from juvenile (p13-15) mouse cortex by applying papain treatment and cytospin centrifugation to attach the cells to a slide. The cells and their complete processes including the PAPs is thus projected in 2D. The entire procedure takes 2.5-3 h. We show by morphometry that the diameter of DIMIGs, including the PAPs is similar to that of astrocytes in situ. In contrast to cell culture, results derived from this procedure allow for direct conclusions relating to (1) the presence of an antigen in cortical astrocytes, (2) subcellular antigen distribution, in particular when localized in the PAPs. The detailed resolution is shown in an exemplary study of the organization of the astrocytic cytoskeleton components actin, ezrin, tubulin, and GFAP. The distribution of connexin 43 in relation to a single astrocyte's process tree is also investigated.
... Glial cells are approximately 10 to 50 times more abundant than neurons and modulate the activity of neurons as well as other glial cells and vascular elements (Magistretti and Pellerin, 1999). In brain tissue, astrocytes have been shown to occupy discrete, yet overlapping domains under normal physiological conditions, and to contribute to brain homeostasis by providing a variety of metabolic functions (Hatton, 2002;Wilhelmsson et al., 2006). They supply growth and trophic factors to support the survival and growth of neural cells and also have the ability to buffer extracellular ion concentrations. ...
Two-dimensional (2-D) neural cell culture systems have served as
physiological models for understanding the cellular and molecular events
that underlie responses to physical and chemical stimuli, control
sensory and motor function, and lead to the development of neurological
diseases. However, the development of three-dimensional (3-D) cell
culture systems will be essential for the advancement of experimental
research in a variety of fields including tissue engineering, chemical
transport and delivery, cell growth, and cell-cell communication. In 3-D
cell culture, cells are provided with an environment similar to tissue,
in which they are surrounded on all sides by other cells, structural
molecules and adhesion ligands. Cells grown in 3-D culture systems
display morphologies and functions more similar to those observed in
vivo, and can be cultured in such a way as to recapitulate the
structural organization and biological properties of tissue. This thesis
describes a hydrogel-based culture system, capable of supporting the
growth and function of several neural cell types in 3-D. Alginate
hydrogels were characterized in terms of their biomechanical and
biochemical properties and were functionalized by covalent attachment of
whole proteins and peptide epitopes. Methods were developed for rapid
cross-linking of alginate hydrogels, thus permitting the incorporation
of cells into 3-D scaffolds without adversely affecting cell viability
or function. A variety of neural cell types were tested including
astrocytes, microglia, and neurons. Cells remained viable and functional
for longer than two weeks in culture and displayed process outgrowth in
3-D. Cell constructs were created that varied in cell density, type and
organization, providing experimental flexibility for studying cell
interactions and behavior. In one set of experiments, 3-D
glial-endothelial cell co-cultures were used to model blood-brain
barrier (BBB) structure and function. This co-culture system was
designed for use as a tool to predict the transport and processing that
occurs prior to drug uptake in the central nervous system (CNS), and to
predict BBB permeability. Electrochemical techniques and
immunohistochemistry were used to validate this model and provide
detailed information about cellular organization and function.
Electrochemical impedance spectroscopy (EIS) provided evidence that
endothelial cells cultured in the presence of astrocytes formed tight
junctions capable of occluding the flow of electrical current. In a
second series of experiments, a microglia-astrocyte co-culture system
was developed to assess the effects of glial cells on electrode
impedance recorded from neural prosthetic devices in vitro. Impedance
measurements were compared with confocal images to determine the effects
of glial cell density and cell type on electrode performance. The
results indicate that EIS data can be used to model components of the
reactive cell responses in brain tissue, and that impedance measurements
recorded in vitro can be compared to measurements recorded in vivo.
Taken together, these results demonstrate that alginate hydrogels can be
used for the creation of 3-D neural cell scaffolds, and that such cell
scaffolds can be used to model a variety of three-dimensional neural
tissues in vitro, that cannot be studied in 2-D cultures.
... While some aspects of MePD neuroanatomy are sexually dimorphic in juvenile animals, including regional volume and synaptic organization (Mizukami et al. 1983;Cooke and Woolley 2005), little is known about astrocytes in the juvenile MePD. Because astrocytes play a critical role in modulating the formation, maintenance and efficacy of synapses (Halassa et al. 2007;Ullian et al. 2001;Nishida and Okabe 2007;Hatton 2002), determining when sex differences in astrocyte morphology arise in the MePD is important for understanding the development of this region. To address this issue, we counted and traced astrocytes in the MePD of juvenile male, female, and Tfm male rats using methods we previously applied to the adult MePD (Johnson et al. 2008). ...
Astrocytes in the posterodorsal portion of the medial amygdala (MePD) are sexually dimorphic in adult rats: males have more astrocytes in the right MePD and more elaborate processes in the left MePD than do females. Functional androgen receptors (ARs) are required for masculinization of MePD astrocytes, as these measures are demasculinized in adult genetic males carrying the testicular feminization mutation (Tfm) of the AR gene, which renders AR dysfunctional. We now report that the number of astrocytes is already sexually dimorphic in the right MePD of juvenile 25-day old (P25) rats. Because Tfm males have as many astrocytes as wildtype males at this age, this prepubertal sexual dimorphism is independent of ARs. After P25, astrocyte number increases in the MePD of all groups, but activation of ARs augments this increase in the right MePD, where more astrocytes are added in males than in Tfm males. Consequently, by adulthood, females and Tfm males have equivalent numbers of astrocytes in the right MePD. Sexual dimorphism in astrocyte arbor complexity in the left MePD arises after P25, and is entirely AR-dependent. Thus, masculinization of MePD astrocytes is a result of both AR-independent processes before the juvenile period and AR-dependent processes afterward. J. Comp. Neurol., 2012. © 2012 Wiley Periodicals, Inc.
... Astrocytes were also present as determined by immunocytochemistry for astrocytic marker GFAP (glial fibrillar acidic protein) (Fig. 9A). The presence of astrocytes is important for cellcell interaction as they secrete factors that modulate the synaptic transmission and enhance the formation of neuronal interconnections (Hatton, 2002). ...
Environmental chemicals have a potential impact on children's health as the developing brain is much more vulnerable to injury caused by different classes of chemicals than the adult brain. This vulnerability is partly due to the fact that very complex processes of cell development and maturation take place within a tightly controlled time frame. So different stages of brain development are susceptible to toxic effects at different time points. Additionally the adult brain is well protected against chemicals by the blood brain barrier (BBB) whereas the placenta only partially protects against harmful chemical exposure. Many metals easily cross the placenta and BBB barrier since even after the birth BBB is not entirely differentiated (until about 6 months after birth). Additionally, the susceptibility of infants and children is due to increased exposure, augmented absorption rates, and less efficient ability of defense mechanism in comparison to adults.
... An expanding body of research indicates that astrocytes are not the passive cells as once imagined and have been linked to the release of gonadotropin releasing hormone (Witkin et al., 1997) and formation of synapses (Ullian et al., 2001;Hatton, 2002;Barker and Ullian, 2008). Moreover, in both the hippocampus (Garcia-Segura et al., 1988;Day et al., 1990Day et al., , 1993Leranth et al., 2008) andhypothalamus (McQueen et al., 1990;Chowen et al., 1995;Mong, 1996;Amateau and McCarthy, 2002), astrocytes are sexually dimorphic and responsive to gonadal hormone manipulations. ...
The posterodorsal medial amygdala (MePD) exhibits numerous sex differences including differences in volume and in the number and morphology of neurons and astroctyes. In adulthood, gonadal hormones, including both androgens and estrogens, have been shown to play a role in maintaining the masculine character of many of these sex differences, but whether adult gonadal hormones maintain the increased number and complexity of astrocytes in the male MePD was unknown. To answer this question we examined astrocytes in the MePD of male and female Long Evans rats that were gonadectomized as adults and treated for 30 days with either testosterone or a control treatment. At the end of treatment brains were collected and immunostained for glial fibrillary acidic protein. Stereological analysis revealed that adult androgen levels influenced the number and complexity of astrocytes in the MePD of both sexes, but the specific effects of androgens were different in males and females. However, sex differences in the number and complexity of adult astrocytes persisted even in the absence of gonadal hormones in adulthood, suggesting that androgens also act earlier in life to determine these adult sex differences. Using immunofluorescence and confocal microscopy, we found robust androgen receptor immunostaining in a subpopulation of MePD astrocytes, suggesting that testosterone may act directly on MePD astrocytes to influence their structure and function.
... Among the factors playing a role in such glial interactions in the nervous systems of both insects (Koussa et al. 2010) and vertebrates (Parys et al. 2010) is Ca 2+ signaling via gap junctions, and one indicator of gap junctional communication is the presence of dye coupling as revealed by fluorescent dyes (Hossain et al. 1995; Ball et al. 2007; Lanosa et al. 2008; Koussa et al. 2010). In vertebrates, glia are known to communicate via gap junctions (Zahs and Newman 1997; Newman 2001; Rela et al. 2010) resulting in cellular networks that may comprise astrocytes alone (Theiss and Meller 2002; Houades et al. 2008; Tanaka et al. 2008), astrocytes and oligodendrocytes (Parys et al. 2010), or astrocytes and neurons (Hatton 2002; Schummers et al. 2008 ). In invertebrates, gap junctional communication has been reported between neurons (Wilcox 1994; Alexopolous et al. 2004; Dykes et al. 2004; Ducret et al. 2006) and between glia (e.g. ...
The central complex of the grasshopper (Schistocerca gregaria) brain comprises a modular set of neuropils, which develops after mid-embryogenesis and is functional on hatching. Early in embryogenesis, Repo-positive glia cells are found intermingled among the commissures of the midbrain, but then redistribute as central complex modules become established and, by the end of embryogenesis, envelop all midbrain neuropils. The predominant glia associated with the central body during embryogenesis are glutamine synthetase-/Repo-positive astrocyte-like glia, which direct extensive processes (gliopodia) into and around midbrain neuropils. We used intracellular dye injection in brain slices to ascertain whether such glia are dye-coupled into a communicating cellular network during embryogenesis. Intracellular staining of individual cells located at any one of four sites around the central body revealed a population of dye-coupled cells whose number and spatial distribution were stereotypic for each site and comparable at both 70 and 100% of embryogenesis. Subsequent immunolabeling confirmed these dye-coupled cells to be astrocyte-like glia. The addition of n-heptanol to the bathing saline prevented all dye coupling, consistent with gap junctions linking the glia surrounding the central body. Since dye coupling also occurred in the absence of direct intersomal contacts, it might additionally involve the extensive array of gliopodia, which develop after glia are arrayed around the central body. Collating the data from all injection sites suggests that the developing central body is surrounded by a network of dye-coupled glia, which we speculate may function as a positioning system for the developing neuropils of the central complex.
... Using these specialized contacts astrocytes can also reach the pia mater. Astrocytic processes in the parenchyma are in contact with and/or ensheath distinct neuronal elements, in particular synapses, with the coverage that varies in different regions of the brain (Ventura and Harris 1999;Hatton 2002). Due to strategic positioning at the cross-roads between the vasculature and neurons, astrocytes can mediate the exchange of molecules between these compartments. ...
J. Neurochem. (2012) 121, 4–27.
Neuroglial cells define brain homeostasis and mount defense against pathological insults. Astroglia regulate neurogenesis and development of brain circuits. In the adult brain, astrocytes enter into intimate dynamic relationship with neurons, especially at synaptic sites where they functionally form the tripartite synapse. At these sites, astrocytes regulate ion and neurotransmitter homeostasis, metabolically support neurons and monitor synaptic activity; one of the readouts of the latter manifests in astrocytic intracellular Ca2+ signals. This form of astrocytic excitability can lead to release of chemical transmitters via Ca2+-dependent exocytosis. Once in the extracellular space, gliotransmitters can modulate synaptic plasticity and cause changes in behavior. Besides these physiological tasks, astrocytes are fundamental for progression and outcome of neurological diseases. In Alzheimer’s disease, for example, astrocytes may contribute to the etiology of this disorder. Highly lethal glial-derived tumors use signaling trickery to coerce normal brain cells to assist tumor invasiveness. This review not only sheds new light on the brain operation in health and disease, but also points to many unknowns.
... In literatures, it has been reported that astrocytes mainly function as supporting cells in the central nervous system. In addition, astrocytes are known to release neuronal substances modulating synaptic transmission and hindering harmful pathways [6, 7, 17]. Initially we hypothesized that if neurons were cocultured with astrocytes , the migration of neuronal cell bodies could appear at a time point earlier than 3 weeks by electrical stimulation. ...
Recently, we found that electrical stimulation can induce neuronal migration in neural networks cultured for more than 3 weeks on microelectrode arrays. Immuno-cytochemistry data showed that the aggregation of neurons was related to the emergence of astrocytes in culture. In this study, when neurons were cocultured with astrocytes, electrical stimulation could induce the migration of neuronal cell bodies after only 1 week in culture, while the same stimulation paradigm caused neural necrosis in neuron-only cultures. In addition, the stimulation-induced migration was inhibited by blocking action potentials in neural networks using the voltage-gated sodium channel blocker, tetrodo-toxin. Immunocytochemistry was performed to monitor precisely the neuronal migration and count the number of neurons. These results indicate that neuronal migration of cell bodies is dependent on neuronal activity evoked by electrical stimulation and can be enhanced by coculturing with astrocytes. We believe this method can be employed as a means for modifying neural networks and improving the interface between electrodes and neurons.
... In contrast APJ-ir has been found in the nerve terminals of the rat posterior pituitary gland [51]. Our study in mice suggests that APJ mRNA is present in an unidentified posterior pituitary cell type that may be resident pituicytes or glial cells where other GPCRs including the V1a receptor are known to be expressed and speculated to indirectly influence neurohypophysial hormone release [15]. The extent of APJ binding sites, i.e. widespread rather than restricted to scattered cells, could suggest that APJ is expressed in both cells and nerve terminals in the mouse posterior pituitary. ...
The G protein-coupled apelin receptor (APJ) binds the endogenous peptide apelin and has been shown to have roles in many physiological systems. Thus far, distribution studies have predominantly been conducted in the rat and there is limited knowledge of the cellular distribution of APJ in mouse or human tissues. As recent functional studies have been conducted in APJ knock-out mice (APJ KO), in this study we undertook to characterize APJ mRNA and I(125)[Pyr(1)]apelin-13 binding site distribution in mouse tissues to enable correlation of distribution with function. We have utilized in situ hybridization histochemistry (ISHH) using APJ riboprobes, which revealed strong hybridization specifically in the paraventricular (PVN) and supraoptic (SON) nuclei of the hypothalamus and in the anterior pituitary, with marginally lower levels in the posterior pituitary. In the periphery, strong hybridization was observed in the lung, heart, adrenal cortex, renal medulla, ovary and uterus. Autoradiographic binding to APJ with I(125)[Pyr(1)]apelin-13 exhibited significant binding in the anterior pituitary, while lower levels were observed in the posterior pituitary and PVN and SON. In the periphery, strong receptor binding was observed in tissues exhibiting intense riboprobe hybridization, indicating a good correlation between receptor transcription and translation. While the distribution of APJ mRNA and functional protein in the mouse shows similarities to that of the rat, we report a species difference in central APJ distribution and in the pituitary gland.
... In rats, these include changes in dopamine, oxytocin, vasopressin and corticotropin releasing hormone neurotransmitter systems [1,2,3,4]. Changes associated with the postpartum period are not limited to neurons, however; astrocytic changes have also been documented in several hypothalamic areas [5]. Theodosis and her colleagues, for example, have documented morphological changes in astrocytes within the supraoptic (SON) and paraventricular (PVN) nuclei of the hypothalamus that have been associated with the bolus release of oxytocin that occurs during milk letdown [6,7,8,9]. ...
An upregulation of the astrocytic proteins GFAP and bFGF within area 2 of the cingulate cortex (Cg2) occurs within 3 hours of parturition in rats. These changes are the result of an interaction between hormonal state and maternal experience and are associated with increased dendritic spine density in this area. Here, we examined whether this upregulation of astrocytic proteins generalized to other glial markers and, in particular those associated with glutamate metabolism. We chose glial markers commonly used to reflect different aspects of glial function: vimentin, like GFAP, is a marker of intermediate filaments; glutamine synthetase (GS), and S-100beta, are used as markers for mature astrocytes and GS has also been used as a specific marker for glutamatergic enzymatic activity. In addition, we examined levels of proteins associated with glutamine synthetase, glutamate, glutamine and two excitatory amino acid transporters found in astrocytes, glt-1 and glast. S100beta immunoreactivity did not vary with reproductive state in either Cg2 or MPOA suggesting no change in the number of mature astrocytes across these conditions. Vimentin-ir did not differ across groups in Cg2, but expression of this protein decreased from Day 1 postpartum onwards in the MPOA. By contrast, GS-ir was increased within 24 h postpartum in Cg2 but not MPOA and similarly to GFAP and bFGF this upregulation of GS resulted from an interaction between hormonal state and maternal experience. Within Cg2, upregulation of GS was not accompanied by changes in the astrocytic glutamatergic transporters, glt-1 and glast, however, an increase in both glutamate and glutamine proteins were observed within the Cg2 of postpartum animals. Together, these changes suggest postpartum upregulation of glutamatergic activity and metabolism within Cg2 that is stimulated by pregnancy hormones and maternal experience.
... Key words: supraoptic nucleus; neurohypophysis; PSA-NCAM; tenascin C; vimentin; S100b; glutamate dehydrogenase Interactions between astrocytes and neurons are essential for several brain functions, including homeostatic and metabolic signalling and trophic and pathophysiological regulation (Hatton, 2002;Barker and Ullian, 2010). The hypothalomo-neurohypophyseal system clearly demonstrates the importance of these functional interactions. ...
The hypothalamo-neurohypophyseal system displays significant plasticity when subjected to physiological stimuli, such as dehydration, parturition, or lactation. This plasticity arises at the neurochemical and electrophysiological levels but also at a structural level. Several studies have demonstrated the role of monoaminergic afferents in controlling neurochemical and electrophysiological plasticity of the supraoptic nucleus (SON) and of the neurohypophysis (NH), but little is known about how the changes in structural plasticity are triggered. We used Tg8 mice, disrupted for the monoamine oxidase A gene, to study monamine involvement in the architecture of the SON and of the NH. SON astrocytes in Tg8 mice displayed an active status, characterized by an increase in S100β expression and a significant decrease in vimentin expression, with no modification in glial fibrillary acidic protein (GFAP) levels. Astrocytes showed a decrease in glutamate dehydrogenase (GDH) levels, whereas glutamine synthetase (GS) levels remained constant, suggesting a reduction in astrocyte glutamate catabolism. Tenascin C and polysialic acid-neural cell adhesion molecule (PSA-NCAM) expressions were also elevated in the SON of Tg8 mice, suggesting an increased capacity for structural remodelling in the SON. In the NH, similar date were obtained with a stability in GFAP expression and an increase in PSA-NCAM immunostaining. These results establish monoamine (serotonin and noradrenaline) involvement in SON and NH structural arrangement. Monoamines therefore appear to be crucial for the coordination of the neurochemical and structural aspects of neuroendocrine plasticity, allowing the hypothalamo-neurohypopyseal system to respond appropriately when stimulated.
... Further, parvocellular and magnocellular neurons of the PVN/ SON coexpress many neuroactive substances (e.g., CRF, galanin, cholecystokinin, enkephalin, and vasoactive intestinal peptide (VIP)) that may have a paracrine/autocrine action on PVN/SON neurons[39,47]. Importantly, in response to various physiological conditions the PVN and SON exhibit a considerable degree of morphological (e.g., glial cell remodeling[115]) and functional plasticity. This can manifest itself by changes in neuronal excitability[84,322]that may be accompanied by enhanced co-expression of certain neuropeptides (e.g., VP in CRF pPVN neurons after stress exposure[2,39]) and/or altered neuro-transmitter/peptide release[42,202]. ...
G protein-coupled receptors (GPCRs) are the largest family of transmembrane receptors in the mammalian genome. They are activated by a multitude of different ligands that elicit rapid intracellular responses to regulate cell function. Unsurprisingly, a large proportion of therapeutic agents target these receptors. The paraventricular nucleus (PVN) and supraoptic nucleus (SON) of the hypothalamus are important mediators in homeostatic control. Many modulators of PVN/SON activity, including neurotransmitters and hormones act via GPCRs--in fact over 100 non-chemosensory GPCRs have been detected in either the PVN or SON. This review provides a comprehensive summary of the expression of GPCRs within the PVN/SON, including data from recent transcriptomic studies that potentially expand the repertoire of GPCRs that may have functional roles in these hypothalamic nuclei. We also present some aspects of the regulation and known roles of GPCRs in PVN/SON, which are likely complemented by the activity of 'orphan' GPCRs.
... Mature astrocytes generally have large arbors extending from their cell bodies, and in the cortex, it has been estimated that each astrocyte connects on average to approximately four neurons, 300–600 dendrites, and 1,000,000 synapses (Halassa et al., 2007). Astrocyte processes are closely involved in the formation of new synapses and these processes are highly mobile and extend or retract to modulate contact between neurons (Hatton, 2002; Nishida and Okabe, 2007; Ullian et al., 2001). Interestingly, increased branch complexity in astrocytes has been associated with an increase in functional synapses in some regions (Elmariah et al., 2005; Pfrieger and Barres, 1997) but has also been associated with decreases in dendritic spines (Mong et al., 2001). ...
The amygdala has received considerable attention because of its established role in specific behaviors and disorders such as anxiety, depression, and autism. Studies have revealed that the amygdala is a complex and dynamic brain region that is highly connected with other areas of the brain. Previous works have focused on neurons, demonstrating that the amygdala in rodents is highly plastic and sexually dimorphic. However, our more recent work explores sex differences in nonneuronal cells, joining a rich literature concerning glia in the amygdala. Prior investigation of glia in the amygdala can generally be divided into disease-related and hormone-related categories, with both areas of research producing interesting findings concerning glia in this important brain region. Despite a wide range of research topics, the collected findings make it clear that glia in the amygdala are sensitive and plastic cells that respond and develop in a highly region specific manner.
Lipids comprise an extremely heterogeneous group of compounds that perform a wide variety of biological functions. Traditional view of lipids as important structural components of the cell and compounds playing a trophic role is currently being supplemented by information on the possible participation of lipids in signaling, not only intracellular, but also intercellular. The review article discusses current data on the role of lipids and their metabolites formed in glial cells (astrocytes, oligodendrocytes, microglia) in communication of these cells with neurons. In addition to metabolic transformations of lipids in each type of glial cells, special attention is paid to the lipid signal molecules (phosphatidic acid, arachidonic acid and its metabolites, cholesterol, etc.) and the possibility of their participation in realization of synaptic plasticity, as well as in other possible mechanisms associated with neuroplasticity. All these new data can significantly expand our knowledge about the regulatory functions of lipids in neuroglial relationships.
Objective
Astrocyte dysfunctions are related to several central nervous system (CNS) pathologies. Transcriptomic profiling of human mRNAs to investigate astrocyte functions may provide the basic molecular-biological data pertaining to the cellular activities of astrocytes.
Methods
Human Primary astrocytes (HPAs) and human neural stem cell line (HB1.F3) were used for differential digital gene analysis. In this study, a massively parallel sequencing platform, next-generation sequencing (NGS), was used to obtain the digital gene expression (DGE) data from HPAs. A comparative analysis of the DGE from HPA and HB1.F3 cells was performed. Sequencing was performed using NGS platform, and subsequently, bioinformatic analyses were implemented to reveal the identity of the pathways, relatively up- or down-regulated in HPA cells.
Results
The top, novel canonical pathways up-regulated in HPA cells than in the HB1.F3 cells were “Cyclins and cell cycle regulation,” “Integrin signaling,” “Regulation of eIF4 and p70S6K signaling,” “Wnt/β-catenin signaling,” “mTOR signaling,” “Aryl hydrocarbon receptor signaling,” “Hippo signaling,” “RhoA signaling,” “Signaling by Rho family GTPases,” and “Glioma signaling” pathways. The down-regulated pathways were “Cell cycle: G1/S checkpoint regulation,” “eIF2 signaling,” “Cell cycle: G2/M DNA damage checkpoint regulation,” “Telomerase signaling,” “RhoGDI signaling,” “NRF2-mediated oxidative stress response,” “ERK/MAPK signaling,” “ATM signaling,” “Pancreatic adenocarcinoma signaling,” “VEGF signaling,” and “Role of CHK proteins in cell cycle checkpoint control” pathways.
Conclusion
This study would be a good reference to understand astrocyte functions at the molecular level, and to develop a diagnostic test, based on the DGE pattern of astrocytes, as a powerful, new clinical tool in many CNS diseases.
Maintenance of hydromineral balance (HB) is an essential condition for life activity at cellular, tissue, organ and system levels. This activity has been considered as a function of the osmotic regulatory system that focuses on hypothalamic vasopressin (VP) neurons, which can reflexively release VP into the brain and blood to meet the demand of HB. Recently, astrocytes have emerged as an essential component of the osmotic regulatory system in addition to functioning as a regulator of the HB at cellular and tissue levels. Astrocytes express all the components of osmoreceptors, including aquaporins, molecules of the extracellular matrix, integrins, and transient receptor potential channels, with an operational dynamic range allowing them to detect and respond to osmotic changes perhaps more efficiently than neurons. The resultant responses, i.e., astroglial morphological and functional plasticity in the supraoptic and paraventricular nuclei, can be conveyed, physically and chemically, to adjacent VP neurons, thereby influencing HB at the system level. In addition, astrocytes, particularly those in the circumventricular organs, are involved not only in VP-mediated osmotic regulation, but also in regulation of other osmolality-modulating hormones, including natriuretic peptides and angiotensin. Thus, astrocytes play a role in local/brain and systemic HB. The adaptive astrocytic reactions to osmotic challenges are associated with signaling events related to the expression of glial fibrillary acidic protein and aquaporin 4 to promote cell survival and repair. However, prolonged osmotic stress can initiate inflammatory and apoptotic signaling processes, leading to glial dysfunction and a variety of brain diseases. Among many diseases of brain injury and hydromineral disorders, cytotoxic and osmotic cerebral edemas are the most common pathological manifestation. Hyponatremia is the most common cause of osmotic cerebral edema. Overly fast correction of hyponatremia could lead to central pontine myelinolysis. In this review, we summarize and analyze the osmosensory functions of astrocytes and their implications in cerebral edema.
The European Food Safety Authority asked the Panel on Plant Protection Products (PPR) and their residues to deliver a Scientific Opinion on the developmental neurotoxicity (DNT) potential of the neonicotinoid insecticides acetamiprid and imidacloprid. An in vitro study (Kimura-Kuroda et al., 2012) suggested that excitation and/or desensitisation of nicotinic acetylcholine receptors (nAChRs) by these compounds might affect developing mammalian nervous systems as occurs with nicotine. To evaluate the DNT potential of acetamiprid and imidacloprid, the PPR Panel scrutinised the open literature, the draft assessments reports and dossiers submitted for approval. The Panel concludes that both compounds may affect neuronal development and function, although several methodological limitations have been identified. Considering the available DNT studies for imidacloprid and acetamiprid, important uncertainties still remain and further in vivo studies following OECD test guideline (TG) 426 are required to robustly characterise a DNT potential and dose-response relationships, particularly for acetamiprid. The Panel considers that current ARfDs may not be protective enough for the possible DNT of acetamiprid and imidacloprid and no reliable conclusion can be drawn as regards the ADI for acetamiprid. More conservative reference values are proposed based on the analysis of the existing toxicological data. However, the current ADI for imidacloprid is considered adequate to protect against its potential developmental neurotoxic effects. Limitations of the in vitro system used by Kimura-Kuroda et al. (2012) prevent its current use as a screening tool in the regulatory arena. The PPR Panel encourages the definition of clear and consistent criteria at EU level to trigger submission of mandatory DNT studies, which could include development of an integrated DNT testing strategy composed of robust, reliable and validated in vitro assays and other alternative methods complementary to the in vivo TG 426 for assessing the DNT potential of substances.
The neurohypophyseal system consists of the neurons in the hypothalamus that synthesize the hormones vasopressin (also called antidiuretic hormone) and oxytocin, their axons that project to the neural lobe of the pituitary, and their terminals in the posterior pituitary. These hormones are hallmarks of mammalian physiology: vasopressin regulates water retention by the kidneys resulting in formation of concentrated urine, thus allowing mammals to live on land; oxytocin causes constriction of the smooth muscles of the uterus and mammary glands, thus allowing mammals to give birth to live young and provide milk to nourish their offspring. In turn, vasopressin secretion is regulated predominantly by receptors that monitor the osmolality of the extracellular fluid as well as blood volume and pressure, whereas oxytocin secretion is stimulated by stretch of the cervix (during birth) and suckling of the young (during lactation).
Synaptic structures of neuronal contacts with pituicytes in the posterior pituitary (PP) were investigated in detail after wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) injection into the rat hypothalamus including the paraventricular, dorsomedial and arcuate nuclei. The injection resulted in heavy labeling of hypothalamic fibers in the PP. In the neuropil of the PP, there were a lot of terminals containing HRP-reaction product (RP) recognized as electron dense lysosomal-like structures. It was of particular interest that the terminals make synaptic contacts with pituicytes. Although the neuronal contacts were characterized by the structure of presynaptic thickening, occasionally two classes could be distinguished - asymmetrical and symmetrical synaptic contacts (Gray's type I and II). These findings provided the morphological evidence for tight structural relationship of neurons with pituicytes in the PP, as well as neuro-neuronal connections in the central nervous system, and additionally for pituicyte modulation of neurohormone output.
One of the first neuropeptides characterized, the hypothalamo-neurohypophysial hormone oxytocin is essential for milk ejection from the mammary gland during lactation. In response to suckling, oxytocin release is pulsatile, maximizing its ability to contract the myoepithelium, and forming the classic milk ejection reflex. Neurophysiological investigations reveal that this periodicity is underlain by the brief, synchronous bursts of oxytocin neurons in the supraoptic and paraventricular nucleus. This activity is prominent during lactation, is dependent upon the sensory stimulation of the young, is regulated by the somatodendritic release of oxytocin within these two nuclei, and is further dependent upon the neurotransmitters glutamate and noradrenaline. Extensive bilateral, intrahypothalamic communication is essential for full expression of the reflex. There is remarkable neuroplasticity at many levels of this system that begins in late pregnancy and extends during lactation to shape the system for maximal periodic release. This chapter explores the neuronal pathways essential to carry and modulate the reflex, and the neurophysiological mechanisms underlying oxytocin neuronal activation.
The present views of the pathogenesis of neuronal dysfunction in critical conditions are analyzed, by taking into account of impairments of cellular NAD+ metabolism, the activity of NAD+-converting enzymes, including ADP-ribosyl cyclase/CD38, the possibilities of developing new neuroprotective strategies. Key words: neuronal dysfunction, ADP-rybosyl cyclase/CD38, NAD+, critical condition.
Mammalian sleep is regulated by two distinct mechanisms. A circadian oscillator provides timing signals that organize sleep and wake across the 24hour day. A homeostatic mechanism increases sleep drive and sleep amounts (or intensity) as a function of prior time awake. The cellular mechanisms of sleep homeostasis are poorly defined, but are thought to be primarily neuronal. According to one view, sleep homeostasis arises from interactions between subcortical neurons that register sleep pressure and other neurons that promote either sleep or wakefulness. Alternatively, sleep drive may arise independently among neurons throughout the brain in a use-dependent fashion. Implicit in both views is the idea that sleep homeostasis is solely the product of neurons. In this article, I discuss an emerging view that glial astrocytes may play an essential role in sleep homeostasis.
We studied changes in the expression and activity of a bifunctional enzyme, CD38, in neurons and glia induced by perinatal
hypoxia-ischemia in rats. These changes influenced reactive gliosis and functional coupling between ADP-ribosyl cyclase and
connexin 43 in the development of neuron-glia interactions.
Inflammation may be involved in the pathogenesis of Alzheimer's disease (AD). There has been little success with anti-inflammatory drugs in AD, while the promise of anti-inflammatory treatment is more evident in experimental models. A new anti-inflammatory strategy requires a better understanding of molecular mechanisms. Among the plethora of signaling pathways activated by β-amyloid (Aβ) peptides, the nuclear factor-kappa B (NF-κB) pathway could be an interesting target. In virus-infected cells, double-stranded RNA-dependent protein kinase (PKR) controls the NF-κB signaling pathway. It is well-known that PKR is activated in AD. This led us to study the effect of a specific inhibitor of PKR on the Aβ42-induced inflammatory response in primary mixed murine co-cultures, allowing interactions between neurons, astrocytes and microglia.
Primary mixed murine co-cultures were prepared in three steps: a primary culture of astrocytes and microglia for 14 days, then a primary culture of neurons and astrocytes which were cultured with microglia purified from the first culture. Before exposure to Aβ neurotoxicity (72 h), co-cultures were treated with compound C16, a specific inhibitor of PKR. Levels of tumor necrosis factor-α (TNFα), interleukin (IL)-1β, and IL-6 were assessed by ELISA. Levels of PT451-PKR and activation of IκB, NF-κB and caspase-3 were assessed by western blotting. Apoptosis was also followed using annexin V-FITC immunostaining kit. Subcellular distribution of PT451-PKR was assessed by confocal immunofluorescence and morphological structure of cells by scanning electron microscopy. Data were analysed using one-way ANOVA followed by a Newman-Keuls' post hoc test
In these co-cultures, PKR inhibition prevented Aβ42-induced activation of IκB and NF-κB, strongly decreased production and release of tumor necrosis factor (TNFα) and interleukin (IL)-1β, and limited apoptosis.
In spite of the complexity of the innate immune response, PKR inhibition could be an interesting anti-inflammatory strategy in AD.
The response to hyperosmotic stresses in the abdominal cavity is regulated, in part, by vasopressin (VP)-secreting neurons in the supraoptic nucleus (SON). How osmotic stress signals are transmitted to the brain is incompletely understood, and whether the transmission routes for osmotic stress signals differ between acute and chronic stresses is unknown. Here we investigated the role of the vagus, splanchnic nerves, and astrocytes in the SON in transducing acute hyperosmotic-stress signals from the abdominal cavity. We found that acute administration of hyperosmotic saline triggered the activation of neurons as well as astrocytes in the SON and the adjoining ventral glia limitans (SON-VGL). Severing the subdiaphragmatic vagal nerve (SDV) prevented the normal response of cells in the SON to HS treatment and attenuated the release of VP into the bloodstream. Lesioning the splanchnic nerves (SNL) diminished HS-induced release of VP, but to a much lesser extent than SDV. Furthermore, SNL did not significantly affect the up-regulation of Fos in SON neurons or the up-regulation of Fos and GFAP in SON and SON-VGL astrocytes that normally occurred in response to HS and did not affect HS-induced expansion of the SON-VGL. Inhibiting astrocytes with fluorocitrate (FCA) prevented the response of the SON to HS and attenuated the release of VP, similarly to SDV surgery. These results suggest that the vagus is the principle route for the transmission of hyperosmotic signals to the brain and that astrocytes in the SON region are necessary for the activation of SON neurons and the release of VP into the bloodstream.
Activity-dependent modulation of excitable responses from neurohypophysial axons and their secretory swellings has long been recognized as an important regulator of arginine vasopressin and oxytocin release during patterned stimulation. Various activity-dependent mechanisms, including action potential broadening, potassium accumulation, and autocrine or paracrine feedback, have been proposed as underlying mechanisms. However, the relevance of any specific mechanism on net excitability in the intact preparation, during different levels of overall activation, and during realistic stimulation with trains of action potentials has remained largely undetermined. Using high-speed optical recordings and potentiometric dyes, we have quantified the dynamics of global excitability under physiologically more realistic conditions, that is in the intact neurohypophysis during trains of stimuli at varying frequencies and levels of overall activity. Net excitability facilitated during stimulation at low frequencies or at low activity. During persistent high-intensity or high-frequency stimulation, net excitability became severely depressed. Depression of excitable responses was strongly affected by manipulations of extracellular potassium levels, including changes to resting [K(+)](out), increases of interstitial spaces with hypertonic solutions and inhibition of Na(+)/K(+) ATPase activity. Application of the GABA(A) receptor blocker bicuculline or manipulations of Ca(2+) influx showed little effect. Numerical simulation of K(+) accumulation on action potentials of individual axons reproduced optically recorded population responses, including the overall depression of action potential (AP) amplitudes, modest AP broadening and the prominent loss of hyperpolarizing undershoots. Hence, extracellular potassium accumulation dominates activity-dependent depression of neurohypophysial excitability under elevated stimulation conditions. The intricate dependence on the short-term stimulation history and its resulting feedback on neurohypophysial excitability renders [K(+)](out) accumulation a surprisingly complex mechanism for regulating axonal excitability and subsequent neuroendocrine release.
Protoplasmic astrocytes are increasingly thought to interact extensively with neuronal elements in the brain and to influence their activity. Recent reports have also begun to suggest that physiologically, and perhaps functionally, diverse forms of these cells may be present in the CNS. Our current understanding of astrocyte form and distribution is based predominately on studies that used the astrocytic marker glial fibrillary acidic protein (GFAP) and on studies using metal-impregnation techniques. The prevalent opinion, based on studies using these methods, is that astrocytic processes overlap extensively and primarily share the underlying neuropil. However, both of these techniques have serious shortcomings for visualizing the interactions among these structurally complex cells. In the present study, intracellular injection combined with immunohistochemistry for GFAP show that GFAP delineates only ∼15% of the total volume of the astrocyte. As a result, GFAP-based images have led to incorrect conclusions regarding the interaction of processes of neighboring astrocytes. To investigate these interactions in detail, groups of adjacent protoplasmic astrocytes in the CA1 stratum radiatum were injected with fluorescent intracellular tracers of distinctive emissive wavelengths and analyzed using three-dimensional (3D) confocal analysis and electron microscopy. Our findings show that protoplasmic astrocytes establish primarily exclusive territories. The knowledge of how the complex morphology of protoplasmic astrocytes affects their 3D relationships with other astrocytes, oligodendroglia, neurons, and vasculature of the brain should have important implications for our understanding of nervous system function.
Lactation is associated with complex changes of the hypothalamo-neurohypophysial system, and oxytocin released within the hypothalamic supraoptic (SON) and paraventricular nuclei may serve as a signal of communication between the magnocellular nuclei in lactating rats. In the first study, the intranuclear and peripheral release patterns of oxytocin and vasopressin in response to intraperitoneal hypertonic saline were studied in virgin and lactating rats to determine if the reduced osmoresponsiveness of the oxytocinergic and vasopressinergic systems during lactation is reflected by reduced release not only into blood, but also within the SON. Simultaneous microdialysis was performed within the SON and the jugular vein before and up to 6 hr after peripheral osmotic stimulation (3.0 M NaCl, 0.6 ml/100 gm body weight, i.p.). There was an immediate increase in secretion of both oxytocin and vasopressin into blood, whereas peptide release within the SON was delayed and peaked after 4-5 hr. Peripheral release of both peptides was significantly reduced in lactating animals, whereas within the SON release of oxytocin, but not vasopressin, was significantly reduced during lactation. In the second study, cross talk between the SONs--another phenomenon which seems to be characteristic for lactation--was studied. Microdialysis of one SON with hypertonic perfusion medium (with 1 M NaCl) significantly increased the release of oxytocin, vasopressin, and various amino acids (aspartate, glutamate, serine, glutamine, gamma amino butyric acid, and arginine) within the ipsilateral SON. In contrast to virgin female and male animals, this unilateral stimulation of the SON resulted in a transiently increased release of oxytocin in the contralateral SON of lactating rats. The release of vasopressin and amino acids within the contralateral SON of lactating rats remained unchanged, indicating specific activation of contralateral oxytocinergic neurons.
1. To evaluate the implication of taurine in the physiology of supraoptic neurones, we (i) investigated the agonist properties of taurine on glycine and GABAA receptors of supraoptic magnocellular neurones acutely dissociated from adult rats, using whole-cell voltage clamp, (ii) studied the effects of taurine and strychnine in vivo by extracellular recordings of supraoptic vasopressin neurones in anaesthetized rats, and (iii) measured the osmolarity-dependent release of endogenous taurine from isolated supraoptic nuclei by HPLC. 2. GABA, glycine and taurine evoked rapidly activating currents that all reversed close to the equilibrium potential for Cl-, indicating activation of Cl(-)-selective channels. Glycine-activated currents were reversibly blocked by strychnine (IC50 of 35 nM with 100 microM glycine), but were unaffected by the GABAA antagonist gabazine (1-3 microM). GABA-activated currents were reversibly antagonized by 3 microM gabazine, but not by strychnine (up to 1 microM). 3. Responses to 1 mM taurine were blocked by strychnine but not by gabazine and showed no additivity with glycine-induced currents, indicating selective activation of glycine receptors. Responses to 10 mM taurine were partially antagonized by gabazine, the residual current being blocked by strychnine. Thus, taurine is also a weak agonist of GABAA receptors. 4. In the presence of gabazine, taurine activated glycine receptors with an EC50 of 406 microM. Taurine activated at most 70% of maximal glycine currents, suggesting that it is a partial agonist of glycine receptors. 5. In vivo, locally applied strychnine (300 nM) increased and taurine (1 mM) decreased the basal electrical activity of vasopressin neurones in normally hydrated rats. The effect of strychnine was markedly more pronounced in water-loaded rats. 6. Taurine, which is concentrated in supraoptic glial cells, could be released from isolated supraoptic nuclei upon hyposmotic stimulation. Decreases in osmolarity of 15 and 30% specifically enhanced basal release of taurine by 42 and 124%, respectively. 7. We conclude that supraoptic neurones express high amounts of glycine receptors, of which taurine may be regarded as a major natural agonist. We postulate that taurine, which can be released in hyposmotic situations, acts on glycine receptors to exert an inhibitory control on magnocellular neurones during alterations of body fluid homeostasis, implicating an active participation of glial cells in this neuroendocrine regulatory loop.
Mature oxytocin (OT) and vasopressin (AVP) magnocellular neurons of the hypothalamic supraoptic nuclei (SON) autocontrol their electrical activity via somatodendritic release of their respective peptides. Because OT and AVP are synthesized early in development and could play an important role in the maturation of these neurons, we checked whether the peptides are released within the SON and act on their secreting neurons during 3 weeks of postnatal development. We used patch-clamp recordings from SON neurons in rat hypothalamic horizontal slices to show that the spontaneous electrical activity of immature SON neurons is blocked by OT or AVP receptor antagonists, demonstrating a basal somatodendritic release of the peptides. Application of OT or AVP depolarizes SON neurons and stimulates activity typical of the corresponding mature neurons. This effect is directly on SON neurons because it is recorded in dissociated neurons. Radioimmunoassays from isolated SON were used to show that each peptide facilitates its own release at a somatodendritic level, exhibiting a self-sustaining positive feedback loop. This autocontrol is not uniform during development because the proportion of neurons depolarized by the peptides, the amplitude of the depolarization, and the propensity of the peptides to facilitate their own release are maximal during the second postnatal week and decrease thereafter. These data are consistent with a role of autocontrol in the maturation of SON neurons because it is maximal during the delimited period of postnatal development that corresponds to the period of major synapse formation.
Analysis of excitatory synaptic transmission in the rat hypothalamic supraoptic nucleus revealed that glutamate clearance and, as a consequence, glutamate concentration and diffusion in the extracellular space, is associated with the degree of astrocytic coverage of its neurons. Reduction in glutamate clearance, whether induced pharmacologically or associated with a relative decrease of glial coverage in the vicinity of synapses, affected transmitter release through modulation of presynaptic metabotropic glutamate receptors. Astrocytic wrapping of neurons, therefore, contributes to the regulation of synaptic efficacy in the central nervous system.
Magnocellular neurons located in the supraoptic nucleus send their principal axons to terminate in the neurohypophysis, where they release vasopressin and oxytocin into the blood circulation. This magnocellular hypothalamo-neurohypophysial system is known to undergo dramatic activity-dependent structural plasticity during chronic physiological stimulation, such as dehydration and lactation. This structural plasticity is accompanied not only by synaptic remodeling, increased direct neuronal membrane apposition, and dendritic bundling in the supraoptic nucleus, but also organization of neurovascular contacts in the neurohypophysis. The adjacent glial cells actively participate in these plastic changes in addition to magnocellular neurons themselves. Many molecules that are possibly concerned with dynamic structural remodeling are highly expressed in the hypothalamo-neurohypophysial system, although they are generally at low expression levels in other regions of adult brains. Interestingly, some of them are highly expressed only in embryonic brains. On the basis of function, these molecules are classified mainly into two categories. Cytoskeletal proteins, such as tubulin, microtubule-associated proteins, and intermediate filament proteins, are responsible for changing both glial and neuronal morphology and location. Cell adhesion molecules, belonging to immunoglobulin superfamily proteins and extracellular matrix glycoproteins, also participate in neuronal-glial, neuronal-neuronal, and glial-glial recognition and guidance. Thus, the hypothalamo-neurohypophysial system is an interesting model for elucidating physiological significance and molecular mechanisms of activity-dependent structural plasticity in adult brains.
Activity of the magnocellular neurons that synthesize vasopressin in the supraoptic and paraventricular nuclei of the hypothalamus is modulated by local release of the neuropeptide within the nuclei. V1a and V1b vasopressin receptor genes are expressed in these cells. The present study reports the localization of V1a and V1b receptors using multiple labeling immunocytochemistry. Both receptors are mainly located in vasopressinergic magnocellular neurons and colocalized with vasopressin in cytoplasmic vesicles dispersed throughout the cell. Possible functional modifications of the mRNA and protein levels of the V1a receptor, the major isoform, were also investigated by semiquantitative in situ hybridization and immunocytochemistry in rats submitted to reduced or increased water intake. V1a mRNA and receptor levels varied with water balance. V1a mRNA level dropped in rats submitted to high water intake. Conversely, dehydration up-regulated the V1a receptor content. These observations suggest that the pathways that regulate the expression of the genes encoding vasopressin and the V1a receptor are linked, which fits the present findings that the two partners are colocalized in cytoplasmic vesicles. Colocalization might explain how V1 autoreceptors are controlled by cell activity and/or local concentration of vasopressin (released locally by the neurons themselves), allowing fine adjustment of magnocellular neuron activity.
Arginine vasopressin- (AVP) and oxytocin- (OXT) secreting magnocellular neurons undergo gross structural changes with chronic physiological stimulation. Here, we investigated subcellular aspects of plasticity in rat neurohypophysial terminals during dehydration. Ultrastructural analyses demonstrated that chronic dehydration by 2% NaCl drinking for 7 days significantly decreased the numbers of neurosecretory granules and microvesicles but not the numbers of mitochondria. Moreover, in dehydrated rats, terminals making neurovascular contacts enlarged, whereas terminals in apposition to astrocytes, i.e., neuroglial contacts, became smaller. Western blot analyses demonstrated significant decreases in the levels of F3 and Thy-1 together with those of AVP- and OXT-neurophysin, but the levels of synaptophysin, SNAP-25, and GAP-43 were unchanged. Both F3 and Thy-1 were recovered in the buffer-insoluble pellet, and phosphatidyl inositol-specific phospholipase C treatment released both molecules from the crude membrane fraction, indicating that they are attached to terminal membranes by glycosylphosphatidyl inositol anchors. Confocal microscopic observations demonstrated that F3 colocalized with Thy-1 in the same terminals of magnocellular neurons. In contrast, the level of calretinin, a Ca2+ binding protein was significantly increased with chronic dehydration. Thus, the present results suggest that enhancement of neurovascular contacts results from rearrangement of terminal-astrocyte and terminal-vessel contacts rather than enlargement or sprouting of magnocellular terminals themselves. The down-regulation of F3 and Thy-1 may contribute to enhancement of neurovascular contacts that accompany increased peptide release during dehydration. J. Comp. Neurol. 434:413–427, 2001. © 2001 Wiley-Liss, Inc.
Taurine, a γ-aminobutyric acid (GABA)-like acidic amino acid, has previously been shown to be prominently localized to astrocytes in the supraoptic nucleus, the neurons of which contain only small amounts, and to have inhibitory actions on supraoptic neuronal activity. In the present study, taurine distribution in the neurohypophysis was determined by using a well-characterized monoclonal antibody against taurine itself. Preembedding immunohistochemistry was performed at light and electron microscopic levels by using diaminobenzidine and gold-substituted silver-intensified peroxidase (GSSP) methods. At the light microscopic level, the distribution pattern and cellular localization of taurine immunoreactivity corresponded to that of glial fibrillary acidic protein. Pituicyte cell bodies and processes displayed dense taurine immunoreactivity. Electron microscopic observations revealed strong taurine GSSP reactions in these neural lobe astrocytes, but weak taurine reactivity was seen within only some neurosecretory axons. High-performance liquid chromatography analyses demonstrated that in vitro hypoosmotic stimulation (reduction of 40 mOsm/kg) of isolated posterior pituitaries resulted in preferential increases in taurine release into the bathing medium without increased release of other amino acids. Conversely, tissue concentrations of taurine significantly decreased with hypoosmotic perfusion, while glutamate, glutamine, and GABA concentrations were not reduced. These results indicate that taurine is mainly concentrated in neurohypophysial astrocytes, which are known to engulf the neurosecretory axonal processes and terminals. Taurine released from pituicytes under basal and hypoosmotic conditions may act to suppress axon terminal depolarization and thereby depress release of neurohypophysial peptides.J.Comp. Neurol. 381:513-523, 1997. © 1997 Wiley-Liss, Inc.
The recent discovery that the neurotransmitter glutamate can trigger actively propagating Ca2+ waves in the cytoplasm of cultured astrocytes suggests the possibility that synaptically released glutamate may trigger similar Ca2+ waves in brain astrocytes in situ. To explore this possibility, we used confocal microscopy and the Ca2+ indicator fluo-3 to study organotypically cultured slices of rat hippocampus, where astrocytic and neuronal networks are intermingled in their normal tissue relationships. We find that astrocytic Ca2+ waves are present under these circumstances and that these waves can be triggered by the firing of glutamatergic neuronal afferents with latencies as short as 2 s. The Ca2+ waves closely resemble those previously observed in cultured astrocytes: they propagate both within and between astrocytes at velocities of 7-27 microns/s at 21 degrees C. The ability of tissue astrocyte networks to respond to neuronal network activity suggests that astrocytes may have a much more dynamic and active role in brain function than has been generally recognized.
An extremely close association exists between the membranes of the neurosecretory endings and the resident astrocytes (pituicytes) of the neurohypophysis. Indeed, synaptoid contacts involving neurosecretory vesicle-containing axons contacting pituicytes have been observed, suggesting pituicytes as targets of the products released from neurosecretory axons. We have investigated the effects of various neural lobe peptides on pituicytes in primary culture from adult neurohypophyses. Using Fura-2 loaded cells and dynamic ratio imaging, we have determined that arginine vasopressin (AVP) or V1- but not V2-receptor agonists, mobilise pituicyte intracellular Ca2+ ([Ca2+]i) in the absence of extracellular Ca2+. AVP was consistently effective at concentrations of 10 nM or higher in elevating [Ca2+]i by 200-1000 nM. These responses could be blocked by V1-antagonists and were shown to be associated with accumulation of phosphoinositides. Oxytocin was also found to mobilise [Ca2+]i but was effective only at higher concentrations than for AVP. Oxytocin-evoked [Ca2+]i elevations were also blocked by V1-antagonists. Raising [K+]0 was ineffective in changing [Ca2+]i suggesting that these cells lack voltage-gated Ca2+ channels. We conclude that pituicytes possess V1-receptors, activation of which mobilises [Ca2+]i, possibly functioning to initiate a Ca(2+)-activated K+ conductance which could contribute to further depolarisation of secretory terminals and facilitate exocytosis.
Neurohypophysial astrocytes, the pituicytes, are known to undergo morphological changes in vivo in response to stimuli that increase the demand for hormone secretion. Similar changes have been induced by beta-adrenergic stimulation both in the isolated, but otherwise intact, neural lobe and in pituicytes cultured from adult rats. Since the predominant beta-receptor subtype in the neural lobe is beta 2, we investigated the possibility that beta 2-receptor activation is mainly responsible for the observed pituicyte responses. In one experiment, cultured pituicytes were induced by noradrenalin to change from flattened amorphous to stellate morphology. Addition of the beta 2-antagonist IPS 339, but not the beta 1-antagonist practolol, significantly reduced (by 30-60%) the number of cells transformed by noradrenalin. In a second experiment, adrenalin, by definition a more potent beta 2-agonist, transformed significantly more pituicytes into stellate shapes than did noradrenalin at the same concentrations (100% vs. 60% increase, respectively). These results support the idea that beta 2-adrenergic receptors are involved in neurohypophysial plasticity. Also, since the neural lobe is outside of the bloodbrain barrier, these findings suggest that adrenal catecholamines participate in altering pituicyte morphology.
Explants of adult rat neurohypophysis were maintained in culture for 14 days. The majority of cells present in the outgrowth of such cultures were identified as pituicytes on the basis of immunostaining for glial fibrillary acidic protein. Pituicytes were also stained by antisera to the membrane glycoprotein antigen Thy-1 and the extracellular matrix glycoprotein fibronectin. The cultures contained naloxone sensitive binding sites for the opioid receptor ligand [3H] dynorphin A 1-8 and peripheral-type benzodiazepine binding sites. Dynorphin binding was visualised over pituicytes following autoradiography. The morphology of cultured pituicytes was regulated by beta-adrenergic receptors present on the cells which, when activated, stimulated rapid transformation from a flattened irregular morphology to a stellate, process-bearing morphology. Dynorphin was without effect on the morphology of cultured pituicytes. These findings are discussed in the context of the known morphological plasticity of pituicytes in vivo.
Exocytosis of neurosecretory granules from dendrites of magnocellular neurons can be visualized electron microscopically after incubation of hypothalamic brain slices in media containing 1.2 mM tannic acid, which stabilizes extracellular peptidergic granule cores, and permits their immunocytochemical identification. Morphometric analysis of stimulated slices demonstrates that exocytosis of neurosecretory granules from the dendrites of magnocellular neurons can account for the vasopressin and oxytocin known to be released into the hypothalamus. Exocytosis from cell bodies of magnocellular neurons was not observed in stimulated slices from normal rats but, when granules had been caused to accumulate in the neuronal somata by prior administration of colchicine, exocytosis of granules from the somata was unambiguously identified. These data demonstrate exocytosis from dendrites and cell bodies of a mammalian peptidergic neuron, and show that all parts of their plasmalemma are competent for exocytosis of granules.
The posterior lobe of the hypophysis consists of neurosecretory nerve fibres and glial cells, both abutting on the perivascular space of sinusoidal capillaries. Nerve fiberes and glial cells are connected with each other by a high number of synaptoid contacts.
Several ultrastructural changes following dehydration have been described by other authors. This study presents the data of some morphometric investigations on normal and dehydrated rats. The relative extent of neuro-vascular contacts is not contstant; it increases significantly from 52 (±2.8)% in normal rats to 67(±1.2)% in rats with a dehydration time of 3 days. Correspondingly the extent of glio-vascular contacts is diminished.
In addition, water-deprived animals show a clear increase in the number of neuro-glial synaptoid contacts with 3.1 (±0.4) per square unit nervous tissue compared to 2.1 (±0.3) per Square unit in untreated controls.
These results indicate a dynamic motility of neuronal and glial elements depending on functional conditions, as was found for the external layer of the median eminence. Furthermore there seems to be a relationship between secretory activity and the number of neuroglial synaptoid contacts.
The release of endogenous oxytocin and vasopressin by rat paraventricular and supraoptic nuclei in vitro during a 10-min period, 30 min after beginning the incubation, was measured radioimmunologically. Mean basal hormone release per 10 min and per pair of nuclei was: 128·4 ± 12·4 ( s.e.m. ) pg vasopressin ( n = 15) and 39·0 ± 3·0 pg oxytocin ( n = 66) for supraoptic nuclei from male rats; 273·9 ± 42·6 pg vasopressin ( n = 11) and 34·2 ± 3·5 pg oxytocin ( n = 15) for supraoptic nuclei from lactating rats; 70·0 ± 8·6 pg vasopressin ( n = 52) and 21·8 ± 1·3 pg oxytocin ( n = 68) for paraventricular nuclei from male rats; 59·1 ± 8·6 pg vasopressin ( n = 10) and 27·0 ± 4·6 pg oxytocin ( n = 16) for paraventricular nuclei from lactating rats.
In male and lactating rats, both nuclei contained and released more vasopressin than oxytocin. For oxytocin alone, the paraventricular nucleus of male rats contained and released significantly less hormone than the supraoptic nucleus. This difference was not apparent in lactating rats. For vasopressin alone, the paraventricular nucleus contained and released significantly less hormone than the supraoptic nucleus in both male and lactating rats. When the hormone released was calculated as a percentage of the total tissue content the release was about 0·9% for oxytocin from both nuclei in male and lactating rats and also for vasopressin in lactating rats, but was only about 0·5% for vasopressin from both nuclei in male rats.
The influence of oxytocin and analogues of oxytocin (including one antagonist) upon the release of oxytocin and vasopressin was studied. Adding oxytocin to the incubation medium (0·4–4 nmol/l solution) induced a dose-dependent rise in oxytocin release from both nuclei of male or lactating rats. A 4 nmol/l solution of isotocin had a similar effect to a 0·4 nmol/l solution of oxytocin, but arginine-vasopressin never affected basal release of oxytocin. In no case was vasopressin release modified.
An oxytocin antagonist (1 μmol/l solution) significantly reduced basal oxytocin release and blocked the stimulatory effect normally induced by exogenous oxytocin, as did gallopamil hydrochloride (D600, 10 μmol/l solution), a Ca ²⁺ channel blocker, or incubation in a Ca ²⁺ -free medium.
These findings are discussed in relation to the literature on the central effects of neurohypophysial peptides. It may be concluded that the regulatory role of endogenous oxytocin in the hypothalamus on the milk-ejection reflex could result from its local release in the extracellular spaces of magnocellular nuclei.
J. Endocr. (1984) 102 , 63–72
Taurine is an inhibitory amino acid that hyperpolarizes magnocellular neurosecretory neurons. To determine which cell types in the rat supraoptic nucleus contain taurine, we used a monoclonal antibody raised against a taurine conjugate. Preembedding immunocytochemistry was carried out at the light and electron microscopic levels using diaminobenzidine and gold-substituted silver-intensified peroxidase as markers. We report the presence of taurine in all cellular compartments of the supraoptic nucleus, except axons, with variable labeling intensities among the different compartments. Few cell bodies of magnocellular neurons were immunoreactive, but many distal dendrites and some proximal ones showed weak-to-moderate levels of immunoreactivity. Strong immunoreactivity was found over glial cell bodies and their processes, in particular in the ventral glial lamina of the supraoptic nucleus. Large astrocytic processes labeled with the taurine antibody included the endfeet participating in the glial limitans around capillaries and at the ventral surface of the hypothalamus. Other types of immunoreactive astrocytic profiles were found scattered within the neuropil where these processes participated in different interactions with the neuronal elements of the supraoptic nucleus. Immunoreactive glial expansions, sometimes even the main process of the glial cell, engulfed axonal boutons. Other labeled glial processes were found between two magnocellular perikarya or closely apposed to the membrane of axonal boutons contacting the neuronal cell bodies. The frequent finding of closely apposed glial and dendritic elements bearing different levels of taurine-like immunoreactivity suggests that exchange of taurine between those two compartments may occur. We propose that taurine could be released from supraoptic glia by a small decrease in osmolarity or by receptor-mediated mechanisms during conditions of low hormonal (vasopressin and/or oxytocin) needs. Such released taurine could then act on presynaptic or postsynaptic sites, or both, to exert its neuromodulatory actions.
In the dendritic region of the supraoptic nucleus of young adult rats we found ultrastructural evidence of rapid (< 30 min) morphological reorganization following transcardial perfusion of the anesthetized animals with high (325 mOsm), but not with normal, osmolality (302 mOsm) balanced salt solutions. The changes observed included significant increases in dendritic bundling and increased numbers of multiple axodendritic synapses (where one presynaptic terminal contacted two or more postsynaptic dendrites). In our initial experiments the perfusion medium contained tannic acid, but when the tannic acid was omitted in a separate set of experiments, the results were both similar and more striking. These studies indicate that the plasticity previously seen in this region under physiological conditions can occur rapidly and probably involves astrocytic process withdrawal from between adjacent dendrites. This change in the distribution of astrocytic processes allows previously single synapses access to additional postsynaptic dendritic membrane. Together, increased dendritic bundling and multiple synapses may facilitate the synchronization of cell firing and, thus, hormone release.
During the past two decades several revisions of the concepts underlying interneuronal communication in the central nervous system have been advanced. We propose here to classify communicational phenomena between cells of the central neural tissue under two general frames: "wiring" and "volume" transmission. "Wiring" transmission is defined as intercellular communication occurring through a well-defined connecting structure. Thus, wiring transmission is characterized by the presence of physically identifiable communication channels within the neuronal and/or glial cell network. It includes synaptic transmission but also other types of intercellular communication through a connecting structure (e.g., gap junctions). "Volume" transmission is characterized by signal diffusion in a three-dimensional fashion within the brain extracellular fluid. Thus, multiple, structurally often not well characterized extracellular pathways connect intercommunicating cells. Volume transmission includes short- (but larger than synaptic cleft, i.e. about 20 nm) and long-distance diffusion of signals through the extracellular and cerebrospinal fluid. It must be underlined that the definitions of wiring and volume transmission focus on the modality of transmission and are neutral with respect to the source and target of the transmission, as well as type of informational substance transmitted. Therefore, any cell present in the neural tissue (neurons, astroglia, microglia, ependyma, tanycytes, etc.) can be a source or a target of wiring and volume transmission. In this paper we discuss the basic definitions and some distinctive characteristics of the two types of transmission. In addition, we review the evidence for different types of intercellular communication besides synaptic transmission in the central nervous system during phylogeny, and in vertebrates in physiological and pathological conditions.
Toward the end of pregnancy, mammalian mothers undergo a cascade of hormonal, neural and somatic events that culminate in birth and lactation. Milk ejection, the final step in lactation, is regulated by high-frequency, synchronized firing of oxytocinergic neurons in the magnocellular nuclei of the hypothalamus. This synchronization of neural activity may be regulated by an increase in the cell-to-cell coupling of magnocellular neurons by connexins. Direct intercellular channels between neurons are formed by the connexin 32 protein. The present study examined the pattern of connexin 32 mRNA levels using semiquantitative in situ hybridization histochemistry with isotopically labeled cRNA probes complementary to connexin 32 and connexin 43, found in neurons/oligodendrocytes and astrocytes, respectively. Connexin 32 mRNA levels were relatively low in the supraoptic nucleus of virgin females and were dramatically up-regulated during late pregnancy. Immediately after birth, connexin 32 mRNA levels dropped and were similar to levels in virgins. On the 13th day of lactation when dye-coupling between magnocellular neurons is high, connexin 32 message levels were again very high. Levels of the astrocytic connexin 43 were no different in lactating and virgin animals. These results demonstrate that there are two peaks of connexin 32 expression, one immediately preceding parturition and one during lactation. So far, only the second peak of connexin 32 is known to be related to an increase in dye-coupling of magnocellular neurons in the supraoptic nucleus, suggesting that in this case, the elevation of connexin 32 message levels leads to the subsequent increase in intercellular coupling.
In mammals, the magnocellular neurons of the supraoptic nucleus (SON) have been classified into vasopressin- (VP) and oxytocin- (OT) producing subtypes. The degree to which these neurons have distinguishable characteristics is considered in the present review. Most of the cytoarchitectonic diversity observed in some Golgi studies has yet to be attributed to differences between OT and VP neurons. The predominant SON cell type is a large bipolar neuron with relatively short and simply branching dendrites. Based on intracellular filling, large multipolar neurons probably represent a small subset of neurosecretory cells. Parvicellular multipolar and bipolar neurons may represent interneurons or subsets of neurosecretory cells. Suggestive evidence that axonal origin and spine density may differ between OT and VP neurons remains to be confirmed in rat. Different fiber systems are thought to preferentially innervate VP or OT subgroups, but only rarely have inputs to OT and VP neurons been compared at the ultrastructural level. Potentially selective inputs to OT somata may derive from the raphe system and the nucleus of the solitary tract, whereas the apparent preferential innervation of VP neurons (e.g. from the A1 region of the ventrolateral medulla) is less certain because of the overlapping dendritic fields of OT and VP neurons. Electrophysiologically, OT and VP neurons are best distinguished in vivo by their reaction to gastric, cardiovascular and suckling stimuli. The firing patterns of activated OT and VP neurons often differ, but can transiently appear indistinguishable in vivo and especially in vitro. Classification in vitro without immunochemical labelling may be aided by the presence of phasic bursting (mostly in VP neurons) and by the differential response of these neurons to certain neurochemicals or to stimulation of certain inputs. The membrane properties of OT and VP neurons are generally similar in vitro, but the range of tests has not been extensive. The depolarizing afterpotential is more often exhibited by, but is not exclusive to, VP neurons.
Taurine, a gamma-aminobutyric acid (GABA)-like acidic amino acid, has previously been shown to be prominently localized to astrocytes in the supraoptic nucleus, the neurons of which contain only small amounts, and to have inhibitory actions on supraoptic neuronal activity. In the present study, taurine distribution in the neurohypophysis was determined by using a well-characterized monoclonal antibody against taurine itself. Preembedding immunohistochemistry was performed at light and electron microscopic levels by using diaminobenzidine and gold-substituted silver-intensified peroxidase (GSSP) methods. At the light microscopic level, the distribution pattern and cellular localization of taurine immunoreactivity corresponded to that of glial fibrillary acidic protein. Pituicyte cell bodies and processes displayed dense taurine immunoreactivity. Electron microscopic observations revealed strong taurine GSSP reactions in these neural lobe astrocytes, but weak taurine reactivity was seen within only some neurosecretory axons. High-performance liquid chromatography analyses demonstrated that in vitro hypoosmotic stimulation (reduction of 40 mOsm/kg) of isolated posterior pituitaries resulted in preferential increases in taurine release into the bathing medium without increased release of other amino acids. Conversely, tissue concentrations of taurine significantly decreased with hypoosmotic perfusion, while glutamate, glutamine, and GABA concentrations were not reduced. These results indicate that taurine is mainly concentrated in neurohypophysial astrocytes, which are known to engulf the neurosecretory axonal processes and terminals. Taurine released from pituicytes under basal and hypoosmotic conditions may act to suppress axon terminal depolarization and thereby depress release of neurohypophysial peptides.
1. Taurine, prominently concentrated in glial cells in the supraoptic nucleus (SON), is probably involved in the inhibition of SON vasopressin neurones by peripheral hypotonic stimulus, via activation of neuronal glycine receptors. We report here the properties and origin of the osmolarity-dependent release of preloaded [3H]taurine from isolated whole SO nuclei. 2. Hyposmotic medium induced a rapid, reversible and dose-dependent increase in taurine release. Release showed a high sensitivity to osmotic change, with a significant enhancement with less than a 5% decrease in osmolarity. Hyperosmotic stimulus decreased basal release. 3. Evoked release was independent of extracellular Ca2+ and Na+, and was blocked by the Cl- channel blockers DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid) and DPC (N-phenylanthranilic acid), suggesting a diffusion process through volume-sensitive Cl- channels. 4. Evoked release was transient for large osmotic reductions (> or = 15%), probably reflecting regulatory volume decrease (RVD). However, it was sustained for smaller changes, suggesting that taurine release induced by physiological variations in osmolarity is not linked to RVD. 5. Basal and evoked release were strongly inhibited by an incubation of the tissue with the glia-specific toxin fluorocitrate, but were unaffected by a neurotoxic-treatment with NMDA, demonstrating the glial origin of the release of taurine in the SON. 6. The high osmosensitivity of taurine release suggests an important role in the osmoregulation of the SON function. These results strengthen the notion of an implication of taurine and glial cells in the regulation of the whole-body fluid balance through the modulation of vasopressin release.
Reviewed in this article are those studies that have contributed heavily to our current conceptualizations of glial participation in the functioning of the magnocellular hypothalamo-neurohypophysial system. This system undergoes remarkable morphological and functional reorganization induced by increased demand for peptide synthesis and release, and this reorganization involves the astrocytic elements in primary roles. Under basal conditions, these glia appear to be vested with the responsibility of controlling the neuronal microenvironment in ways that reduce neuronal excitability, restrict access to neuronal membranes by neuroactive substances and deter neuron neuron interactions within the system. With physiological activation, the glial elements, via receptor-mediated mechanisms, take up new positions. This permissively facilitates neuron neuron interactions such as the exposure of neuronal membranes to released peptides and the formation of gap junctions and new synapses, enhances and prolongs the actions of those excitatory neurotransmitters for which there are glial uptake mechanisms, and facilitates the entry of peptides into the blood. In addition, subpopulations of these glia either newly synthesize or increase synthesis of neuroactive peptides for which their neuronal neighbors have receptors. Release of these peptides by the glia or their functional roles in the system have not yet been demonstrated.
Astrocytes can release glutamate in a calcium-dependent manner and consequently signal to adjacent neurons. Whether this glutamate release pathway is used during physiological signaling or is recruited only under pathophysiological conditions is not well defined. One reason for this lack of understanding is the limited knowledge about the levels of calcium necessary to stimulate glutamate release from astrocytes and about how they compare with the range of physiological calcium levels in these cells. We used flash photolysis to raise internal calcium in astrocytes, while monitoring astrocytic calcium levels and glutamate, which evoked slow inward currents that were recorded electrophysiologically from single neurons grown on microislands of astrocytes. With this approach, we demonstrate that modest changes of astrocytic calcium, from 84 to 140 nM, evoke substantial glutamatergic currents in neighboring neurons (-391 pA), with a Hill coefficient of 2.1 to 2.7. Because the agonists glutamate, norepinephrine, and dopamine all raise calcium in astrocytes to levels exceeding 1.8 microM, these quantitative studies demonstrate that the astrocytic glutamate release pathway is engaged at physiological levels of internal calcium. Consequently, the calcium-dependent release of glutamate from astrocytes functions within an appropriate range of astrocytic calcium levels to be used as a signaling pathway within the functional nervous system.
Activity of the magnocellular neurons that synthesize vasopressin in the supraoptic and paraventricular nuclei of the hypothalamus is modulated by local release of the neuropeptide within the nuclei. V(1a) and V(1b) vasopressin receptor genes are expressed in these cells. The present study reports the localization of V(1a) and V(1b) receptors using multiple labeling immunocytochemistry. Both receptors are mainly located in vasopressinergic magnocellular neurons and colocalized with vasopressin in cytoplasmic vesicles dispersed throughout the cell. Possible functional modifications of the mRNA and protein levels of the V(1a) receptor, the major isoform, were also investigated by semiquantitative in situ hybridization and immunocytochemistry in rats submitted to reduced or increased water intake. V(1a) mRNA and receptor levels varied with water balance. V(1a) mRNA level dropped in rats submitted to high water intake. Conversely, dehydration up-regulated the V(1a) receptor content. These observations suggest that the pathways that regulate the expression of the genes encoding vasopressin and the V(1a) receptor are linked, which fits the present findings that the two partners are colocalized in cytoplasmic vesicles. Colocalization might explain how V(1) autoreceptors are controlled by cell activity and/or local concentration of vasopressin (released locally by the neurons themselves), allowing fine adjustment of magnocellular neuron activity.
Oxytocin-secreting neurons of the hypothalamoneurohypophysial system undergo reversible morphological changes whenever they are strongly stimulated. In the hypothalamus, such structural plasticity is represented by modifications in the size and shape of their somata and dendrites, in the extent to which their surfaces are covered by glia, and in the density of their synapses. In the neurohypophysis, there is a parallel reduction in glial (pituicyte) coverage of their axons together, with retraction of pituicyte processes from the perivascular basal lamina and an increase in the number and size of their terminals. These changes occur rapidly, within a few hours. On the other hand, the system returns to its prestimulated condition on arrest of stimulation at a rate that depends on the length of time it has remained activated. Such neuronal–glial changes have several functional consequences. In the hypothalamic nuclei, reduction in astrocytic coverage of oxytocinergic neurons and their synapses modifies extracellular ionic homeostasis and glutamate clearance and, therefore, their overall excitability. Since it results in extensive dendritic bundling, it may also lead to ephaptic interactions and may facilitate dendritic electrotonic coupling. A most important indirect effect may be to permit synaptic remodeling that occurs concomitantly and that results in significant increases in the number of excitatory and inhibitory synapses driving their activity. In the stimulated neurohypophysis, glial retraction results in increased levels of extracellular K+ which can enhance neurohormone release while an enlarged neurovascular contact zone may facilitate diffusion of neurohormone into the circulation.
In the supraoptic nucleus (SON), the incidence of conducting gap junctions (gjs), as indicated by dye coupling, is low in cycling females, but dramatically elevated in nursing mothers. Functionally, this is consistent with the well-established presence of synchronous milk ejection bursts among oxytocin neurons only in the lactating rat. In situ hybridization data, however, revealed elevated gj mRNA expression on the last day of pregnancy, a time when burst firing by putative oxytocin neurons is absent. Using Lucifer Yellow dye coupling, we determined the incidence of high conductance gjs in SONs of proestrous, immediately prepartum, postpartum non-lactating, lactating day 1, and lactating day 9-10 rats. Results indicate that coupling incidence is high only at times when milk ejection bursts are known to occur, and that the elevated gj mRNA expression seen on the last day of pregnancy does not indicate conducting gjs. It is suggested that gj conductance states, but not gj expression, are modulated by plasma estradiol titers.
Towards the middle of the 20th century, neuroanatomy was on the decline. It was revived by the development of two new methods. One was the Nauta-Gygax method, which selectively stained nerve fibers that had been caused to degenerate by experimental lesions. This allowed connections between various parts of the nervous system to be better determined. The second was electron microscopy, which allowed the structure of neurons and the synapses between them to be examined in detail, and eventually this led to a revival of the Golgi impregnation methods. This occurred in the 1970s because of the desire of electron microscopists to determine the origins of the neuronal profiles they encountered in electron micrographs of various parts of the central nervous system. Eventually this led to the development of Golgi/EM techniques, whereby individual impregnated neurons could first be characterized by light microscopy and then thin sectioned for detailed analyses. Examining the axon terminals of such impregnated neurons, especially those in the cerebral cortex, for the first time revealed details of intercellular connections and allowed neuronal circuits to be postulated. However, Golgi/EM had only a brief, but fruitful existence. It was soon superceded by intracellular filling techniques, which allowed the added dimension that the physiological properties of identified neurons could also be determined.
- Vi Kalnins
- L Subrahmanyan
- Opas The Cytoskeleton
- M In
Kalnins VI, Subrahmanyan L, and Opas The cytoskeleton
M. In: Astrocytes: Cell Biology and Pathology of Astrocytes,
edited by Federoff S and Vernadakis A. Orlando, FL: Academic,
1986, p. 27–59.
- V I Kalnins
- L Subrahmanyan
- Opas The Cytoskeleton M
Kalnins VI, Subrahmanyan L, and Opas The cytoskeleton
M. In: Astrocytes: Cell Biology and Pathology of Astrocytes,
edited by Federoff S and Vernadakis A. Orlando, FL: Academic,
1986, p. 27-59.