D J Maxwell

Publications of D J Maxwell

• Organization and neurochemical properties of intersegmental interneurons in the lumbar enlargement of the adult rat.

  Authors: T T Liu, B A Bannatyne, D J Maxwell

  Neuroscience. 12/2010; 171(2):461-84.

  Intersegmental interneurons with relatively short axons perform an important role in the coordination of limb movement but surprisingly little is known about their organization and how theyIntersegmental interneurons with relatively short axons perform an important role in the coordination of limb movement but surprisingly little is known about their organization and how they contribute to neuronal networks in the adult rat. We undertook a series of anatomical tract-tracing studies to label cell bodies and axons of intersegmental neurons in the lumbar cord and characterized their neurochemical properties by using immunocytochemistry. The b-subunit of cholera toxin was injected into L1 or L3 segments of seven rats in the vicinity of lateral or medial motor nuclei. In L5 lumbar segments, cells were found to be concentrated in contralateral lamina VIII, and in ipsilateral lamina VII and laminae V-VI following injections into the lateral and medial motor nuclei respectively. About 25% of labelled cells contained calbindin or calretinin or a combination of both. Calbindin positive cells were mainly distributed within the ipsilateral side of the L5 segment, especially within the ipsilateral dorsal horn whereas there was a concentration of calretinin cells in contralateral lamina VIII. A small population of cells around the central canal were cholinergic. We also examined axon terminals that projected from L1/3 to the L5 contralateral lateral motor nucleus. The majority of these axons were excitatory (75%) and made direct contacts with motoneurons. However, most inhibitory axons in L5 contained a mixture of GABA and glycine (20%) and about 22% of the total population of axons contained calbindin. In contrast, 19% of all intra-segmental axons in the L3 contralateral lateral motor nucleus were found to be purely glycinergic and 17% contained a mixture of GABA and glycine. This study shows that short range interneurons form extensive ipsi- and contralateral projections within the lumbar enlargement and that many of them contain calcium binding proteins. Those projecting contralaterally to motor nuclei are predominantly excitatory.

• Cholinergic terminals in the ventral horn of adult rat and cat: Evidence that glutamate is a cotransmitter at putative interneuron synapses but not at central synapses of motoneurons.

  Authors: T T Liu, B A Bannatyne, E Jankowska, D J Maxwell

  Neuroscience. 04/2009;

  Until recently it was generally accepted that the only neurotransmitter to be released at central synapses of somatic motoneurons was acetylcholine. However, studies on young mice (P0-10) haveUntil recently it was generally accepted that the only neurotransmitter to be released at central synapses of somatic motoneurons was acetylcholine. However, studies on young mice (P0-10) have provided pharmacological evidence indicating that glutamate may act as a cotransmitter with acetylcholine at synapses between motoneurons and Renshaw cells. We performed a series of anatomical experiments on axon collaterals obtained from intracellularly labeled motoneurons from an adult cat and labeled by retrograde transport in adult rats to determine if glutamate is co-localized with acetylcholine by these terminals. We could find no evidence for the presence of vesicular glutamate transporters in motoneuron axon terminals of either species. In addition, we were unable to establish any obvious relationship between motoneuron terminals and the R2 subunit of the AMPA receptor (GluR2). However we did observe a population of cholinergic terminals in lamina VII which did not originate from motoneurons but were immunoreactive for the vesicular glutamate transporter 2 and formed appositions to GluR2 subunits. These were smaller than motoneuron terminals and, unlike them, formed no relationship with Renshaw cells. The evidence suggests that glutamate does not act as a cotransmitter with acetylcholine at central synapses of motoneurons in the adult cat and rat. However, glutamate is present in a population of cholinergic terminals which probably originate from interneurons where its action is via an AMPA receptor.

• Premotor interneurones contributing to actions of feline pyramidal tract neurones on ipsilateral hindlimb motoneurones.

  Authors: K Stecina, E Jankowska, A Cabaj, L G Pettersson, B A Bannatyne, D J Maxwell

  The Journal of physiology. 02/2008; 586(2):557-74.

  The aim of the study was to analyse the potential contribution of excitatory and inhibitory premotor interneurones in reflex pathways from muscle afferents to actions of pyramidal tract (PT) neuronesThe aim of the study was to analyse the potential contribution of excitatory and inhibitory premotor interneurones in reflex pathways from muscle afferents to actions of pyramidal tract (PT) neurones on ipsilateral hindlimb motoneurones. Disynaptic EPSPs and IPSPs evoked in motoneurones in deeply anaesthetized cats by group Ia, Ib and II muscle afferents were found to be facilitated by stimulation of the ipsilateral, as well as of contralateral, PT. The ipsilateral actions were evoked by either uncrossed or double-crossed pathways. The results show that interneurones mediating reflex actions of muscle afferents may be activated strongly enough by PT stimulation to contribute to movements initiated by ipsilateral PT neurones and that PT actions relayed by them might be enhanced by muscle stretches and/or contractions. However, in some motoneurones disynaptic IPSPs and EPSPs evoked from group Ib or II afferents were depressed by PT stimulation. In order to analyse the basis of this depression, the transmitter content in terminals of 11 intracellularly labelled interneurones excited by PT stimulation was defined immunohistochemically and their axonal projections were reconstructed. The interneurones included 9 glycinergic and 2 glutamatergic neurones. All but one of these neurones were mono- or disynaptically excited by group I and/or II afferents. Several projected to motor nuclei and formed contacts with motoneurones. However, all had terminal projections to areas outside the motor nuclei. Therefore both inhibitory and excitatory interneurones could modulate responses of other premotor interneurones in parallel with direct actions on motoneurones.

• On coupling and decoupling of spinal interneuronal networks.

  Authors: E Jankowska, D J Maxwell, B A Bannatyne

  Archives italiennes de biologie. 12/2007; 145(3-4):235-50.

  This review addresses the question of interrelations between spinal interneuronal networks. On the basis of electrophysiological, pharmacological, morphological and immunohistochemical analysis ofThis review addresses the question of interrelations between spinal interneuronal networks. On the basis of electrophysiological, pharmacological, morphological and immunohistochemical analysis of interneurones mediating various reflex actions from muscle receptors and of reticulospinal neurones a considerable degree of interweaving between networks of these neurones has been established. The coupling has been found to occur at the level of several sites of these networks but the review focuses on two of these sites. The first is between dorsal horn interneurones with group II input and their target ipsilaterally and contralaterally projecting intermediate zone and commissural interneurones. The second is between commissural interneurones with input from reticulospinal neurones and their target interneurones. Several ways of both strengthening and weakening of coupling between various interneuronal networks are also briefly reviewed.

• P boutons in lamina IX of the rodent spinal cord express high levels of glutamic acid decarboxylase-65 and originate from cells in deep medial dorsal horn.

  Authors: D I Hughes, M Mackie, G G Nagy, J S Riddell, D J Maxwell, G Szabó, F Erdélyi, G Veress, P Szucs, M Antal, A J Todd

  Proceedings of the National Academy of Sciences of the United States of America. 07/2005; 102(25):9038-43.

  Presynaptic inhibition of primary muscle spindle (group Ia) afferent terminals in motor nuclei of the spinal cord plays an important role in regulating motor output and is produced by a population ofPresynaptic inhibition of primary muscle spindle (group Ia) afferent terminals in motor nuclei of the spinal cord plays an important role in regulating motor output and is produced by a population of GABAergic axon terminals known as P boutons. Despite extensive investigation, the cells that mediate this control have not yet been identified. In this work, we use immunocytochemistry with confocal microscopy and EM to demonstrate that P boutons can be distinguished from other GABAergic terminals in the ventral horn of rat and mouse spinal cord by their high level of the glutamic acid decarboxylase (GAD) 65 isoform of GAD. By carrying out retrograde labeling from lamina IX in mice that express green fluorescent protein under the control of the GAD65 promoter, we provide evidence that the cells of origin of the P boutons are located in the medial part of laminae V and VI. Our results suggest that P boutons represent the major output of these cells within the ventral horn and are consistent with the view that presynaptic inhibition of proprioceptive afferents is mediated by specific populations of interneurons. They also provide a means of identifying P boutons that will be important in studies of the organization of presynaptic control of Ia afferents.

• Transmitter content, origins and connections of axons in the spinal cord that possess the serotonin (5-hydroxytryptamine) 3 receptor.

  Authors: D Conte, E D Legg, A C McCourt, E Silajdzic, G G Nagy, D J Maxwell

  Neuroscience. 02/2005; 134(1):165-73.

  Recent evidence suggests that serotonin has pronociceptive actions in the spinal cord when it acts through 5-hydroxytryptamine (5-HT)(3) receptors. Cells and axon terminals which are concentrated inRecent evidence suggests that serotonin has pronociceptive actions in the spinal cord when it acts through 5-hydroxytryptamine (5-HT)(3) receptors. Cells and axon terminals which are concentrated in the superficial dorsal horn possess this receptor. We performed a series of immunocytochemical studies with an antibody raised against the 5-HT(3A) subunit in order to address the following questions: 1) Are axons that possess 5-HT(3) receptors excitatory? 2) Are 5-HT(3) receptors present on terminals of myelinated primary afferents? 3) What is the chemical nature of dorsal horn cells that possess 5-HT(3) receptors? 4) Do axons that possess 5-HT(3) receptors target lamina I projection cells? Approximately 45% of 5-HT(3A) immunoreactive boutons were immunoreactive for the vesicular glutamate transporter 2 and almost 80% formed synapse-like associations with GluR2 subunits of the AMPA receptor therefore it is principally glutamatergic axons that possess the receptor. Immunoreactivity was not present on myelinated primary afferent axons labeled with the B-subunit of cholera toxin or those containing the vesicular glutamate transporter 1. Calbindin (which is associated with excitatory interneurons) was found in 44% of 5-HT(3A) immunoreactive cells but other markers for inhibitory and excitatory cells were not present. Lamina I projection cells that possessed the neurokinin-1 receptor were associated with 5-HT(3A) axons but the density of contacts on individual neurons varied considerably. The results suggest that 5-HT(3) receptors are present principally on terminals of excitatory axons, and at least some of these originate from dorsal horn interneurons. The relationship between lamina I projection cells and axons possessing the 5-HT(3) receptor indicates that this receptor has an important role in regulation of ascending nociceptive information.

• Axon terminals possessing alpha2C-adrenergic receptors densely innervate neurons in the rat lateral spinal nucleus which respond to noxious stimulation.

  Authors: M J Olave, D J Maxwell

  Neuroscience. 02/2004; 126(2):391-403.

  The lateral spinal nucleus (LSN) in the rat spinal cord contains projection neurons that are densely innervated by peptidergic varicosities which probably originate from spinal interneurons. TheThe lateral spinal nucleus (LSN) in the rat spinal cord contains projection neurons that are densely innervated by peptidergic varicosities which probably originate from spinal interneurons. The alpha2C-adrenoceptor (alpha2C-AR) is present on axon terminals in this nucleus and therefore norepinephrine is likely to modulate input to LSN neurons. We investigated the involvement of LSN neurons in nociceptive transmission and their relationship with axons that possess alpha2C-ARs. Double-labeling immunostaining experiments showed that alpha2C-ARs are present on axon terminals of excitatory and inhibitory interneurons that frequently contain colocalised peptides. Electron microscopy revealed that alpha2C-AR terminals are presynaptic to dendrites and somata of LSN neurons and predominantly form asymmetric synapses. We retrogradely labeled LSN neurons that project to the caudal ventrolateral medulla and combined this with induction of c-Fos expression by peripheral noxious thermal stimulation along with immunolabelling for the alpha2C-AR and the substance P (neurokinin-1) receptor. This enabled us to identify neuronkinin-1 projection neurons in the LSN that express c-Fos and to determine if such cells receive contacts from alpha2C-AR terminals. The results show that some LSN neurons are activated by noxious stimulation and that this input is likely to be modulated by norepinephrine acting on alpha2C-ARs which are present on axon terminals that are presynaptic to LSN neurons.

• Selective loss of spinal GABAergic or glycinergic neurons is not necessary for development of thermal hyperalgesia in the chronic constriction injury model of neuropathic pain.

  Authors: E Polgár, D I Hughes, J S Riddell, D J Maxwell, Z Puskár, A J Todd

  Pain. 08/2003; 104(1-2):229-39.

  GABA and glycine are inhibitory neurotransmitters used by many neurons in the spinal dorsal horn, and intrathecal administration of GABA(A) and glycine receptor antagonists produces behavioural signsGABA and glycine are inhibitory neurotransmitters used by many neurons in the spinal dorsal horn, and intrathecal administration of GABA(A) and glycine receptor antagonists produces behavioural signs of allodynia, suggesting that these transmitters have an important role in spinal pain mechanisms. Several studies have described a substantial loss of GABA-immunoreactive neurons from the dorsal horn in nerve injury models, and it has been suggested that this may be associated with a loss of inhibition, which contributes to the behavioural signs of neuropathic pain. We have carried out a quantitative stereological analysis of the proportions of neurons in laminae I, II and III of the rat dorsal horn that show GABA- and/or glycine-immunoreactivity 2 weeks after nerve ligation in the chronic constriction injury (CCI) model, as well as in sham-operated and nai;ve animals. At this time, rats that had undergone CCI showed a significant reduction in the latency of withdrawal of the ipsilateral hindpaw to a radiant heat stimulus, suggesting that thermal hyperalgesia had developed. However, we did not observe any change in the proportion of neurons in laminae I-III of the ipsilateral dorsal horn that showed GABA- or glycine-immunoreactivity compared to the contralateral side in these animals, and these proportions did not differ significantly from those seen in sham-operated or nai;ve animals. In addition, we did not see any evidence for alterations of GABA- or glycine-immunostaining in the neuropil of laminae I-III in the animals that had undergone CCI. Our results suggest that significant loss of GABAergic or glycinergic neurons is not necessary for the development of thermal hyperalgesia in the CCI model of neuropathic pain.

• Axon terminals possessing the alpha 2c-adrenergic receptor in the rat dorsal horn are predominantly excitatory.

  Authors: M J Olave, D J Maxwell

  Brain research. 04/2003; 965(1-2):269-73.

  In this study we used confocal microscopy to show that most (83.67%) alpha(2C)-adrenergic receptor (alpha(2C)-AR)-immunoreactive terminals in the superficial dorsal horn contain the vesicularIn this study we used confocal microscopy to show that most (83.67%) alpha(2C)-adrenergic receptor (alpha(2C)-AR)-immunoreactive terminals in the superficial dorsal horn contain the vesicular glutamate transporter 2 and hence are glutamatergic. Few (11.33%) terminals contain glutamic acid decarboxylase (a marker for GABAergic axons) and none were associated with the B subunit of cholera toxin (a marker for myelinated primary afferents) or the vesicular glutamate transporter 1. These data indicate that most dorsal horn axons possessing the alpha(2C)-AR are excitatory and add further support to the suggestion that they originate principally from spinal interneurons.

• Characterisation of axon terminals in the rat dorsal horn that are immunoreactive for serotonin 5-HT3A receptor subunits.

  Authors: D J Maxwell, R Kerr, S Rashid, E Anderson

  Experimental brain research. Experimentelle Hirnforschung. Expérimentation cérébrale. 04/2003; 149(1):114-24.

  Serotonin 5-HT(3) receptors are abundant in the superficial dorsal horn and are likely to have an involvement in processing of nociceptive information. It has been shown previously that 5-HT(3)Serotonin 5-HT(3) receptors are abundant in the superficial dorsal horn and are likely to have an involvement in processing of nociceptive information. It has been shown previously that 5-HT(3) receptors are present on primary afferent terminals and some dorsal horn cells. The primary aim of the present study was to determine what classes of primary afferent possess 5-HT(3)A receptor subunits. We performed a series of double- and triple-labelling immunofluorescence experiments. Subunits were labelled with an anti-peptide antibody and primary afferent axons were identified by the presence of calcitonin gene-related peptide (CGRP) and binding of the lectin IB4. Quantitative confocal microscopic analysis revealed that approximately 10% of axons displaying 5-HT(3)A immunoreactivity were also labelled for CGRP but that only 3% of these fibres bind IB4. We also investigated the relationship between immunoreactivity for the subunit and descending serotoninergic systems, axons originating from inhibitory neurons that contain glutamic acid decarboxylase, and axons of a subpopulation of excitatory neurons that contain neurotensin. None of these types of axon was associated with immunoreactivity for receptor subunits. Ultrastructural studies confirmed that punctate immunoreactive structures observed with the light microscope were axon terminals. These terminals invariably formed asymmetric synaptic junctions with dendritic profiles and often contained a mixture of granular and agranular vesicles. Some terminals formed glomerular-like arrangements. Immunoreactive cells were also examined and were found to contain intense patches of reaction product within the cytoplasm. We conclude that the majority (about 87%) of dorsal horn axons that are immunoreactive for 5-HT(3)A receptor subunits do not originate from the subtypes of primary afferent fibres that bind IB4 or contain CGRP. It is likely that most of these axons have an excitatory action and they may originate from dorsal horn interneurons and/or fine myelinated primary afferent fibres.

• Distribution of and organisation of dorsal horn neuronal cell bodies that possess the muscarinic m2 acetylcholine receptor.

  Authors: W Stewart, D J Maxwell

  Neuroscience. 02/2003; 119(1):121-35.

  Cholinergic systems in the dorsal horn are involved in antinociception but little is known about the organisation of receptors that mediate this process. In this study we examined immunocytochemicalCholinergic systems in the dorsal horn are involved in antinociception but little is known about the organisation of receptors that mediate this process. In this study we examined immunocytochemical properties of dorsal horn neuronal cell bodies that express the m2 muscarinic acetylcholine receptor. Tissue was examined with confocal laser scanning microscopy and quantitative analysis performed. Immunoreactive cells were found throughout the dorsal horn and in lamina X. Quantitative analysis revealed that 22% of neuronal somata in the dorsal horn possess the receptor. The greatest concentration of cells was found in deeper laminae (IV-VI) and around lamina X. A proportion of cholinergic cells (labelled with an antibody against choline acetyltransferase) were immunoreactive for the receptor (approximately, 40% of dorsal horn cells and 44% of lamina X cells). Populations of presumed inhibitory interneurons also displayed immunoreactivity for the receptor. Between 27-34% of cells immunoreactive for GABA, nitric oxide synthase and the somatostatin receptor(2A) expressed the receptor but only 8% of parvalbumin-immunoreactive cells displayed receptor immunoreactivity. Cells labelled with neurotensin, which belong to a subgroup of excitatory neurons, displayed no receptor immunoreactivity. A small number neurokinin-1 receptor-immunoreactive cells in lamina I possessed m2 immunoreactivity but 42% of laminae III/IV neurokinin-1 cells possessed it.This study shows that a significant proportion of cell bodies in the dorsal horn express the muscarinic m2 acetylcholine receptor. The receptor is present on some cholinergic neurons and therefore may function as an autoreceptor. It is associated with inhibitory local circuit neurons and may have a role in the modulation of specific inhibitory systems. It is also found on a proportion of projection cells that possess the neurokinin-1 receptor. This could be the basis of some of the antinociceptive actions of acetylcholine.

• Distribution and colocalisation of glutamate decarboxylase isoforms in the rat spinal cord.

  Authors: M Mackie, D I Hughes, D J Maxwell, N J K Tillakaratne, A J Todd

  Neuroscience. 02/2003; 119(2):461-72.

  The inhibitory neurotransmitter GABA is synthesized by glutamic acid decarboxylase (GAD), and two isoforms of this enzyme exist: GAD65 and GAD67. Immunocytochemical studies of the spinal cord haveThe inhibitory neurotransmitter GABA is synthesized by glutamic acid decarboxylase (GAD), and two isoforms of this enzyme exist: GAD65 and GAD67. Immunocytochemical studies of the spinal cord have shown that whilst both are present in the dorsal horn, GAD67 is the predominant form in the ventral horn. The present study was carried out to determine the pattern of coexistence of the two GAD isoforms in axonal boutons in different laminae of the cord, and also to examine the relation of the GADs to the glycine transporter GLYT2 (a marker for glycinergic axons), since many spinal neurons are thought to use GABA and glycine as co-transmitters.Virtually all GAD-immunoreactive boutons throughout the spinal grey matter were labelled by both GAD65 and GAD67 antibodies; however, the relative intensity of staining with the two antibodies varied considerably. In the ventral horn, most immunoreactive boutons showed much stronger labelling with the GAD67 antibody, and many of these were also GLYT2 immunoreactive. However, clusters of boutons with high levels of GAD65 immunoreactivity were observed in the motor nuclei, and these were not labelled with the GLYT2 antibody. In the dorsal horn, some GAD-immunoreactive boutons had relatively high levels of labelling with either GAD65 or GAD67 antibody, whilst others showed a similar degree of labelling with both antibodies. GLYT2 immunoreactivity was associated with many GAD-immunoreactive boutons; however, this did not appear to be related to the pattern of GAD expression.It has recently been reported that there is selective depletion of GAD65, accompanied by a loss of GABAergic inhibition, in the ipsilateral dorsal horn in rats that have undergone peripheral nerve injuries [J Neurosci 22 (2002) 6724]. Our finding that some boutons in the superficial laminae showed relatively high levels of GAD65 and low levels of GAD67 immunoreactivity is therefore significant, since a reduction in GABA synthesis in these axons may contribute to neuropathic pain.

• The expression of vesicular glutamate transporters VGLUT1 and VGLUT2 in neurochemically defined axonal populations in the rat spinal cord with emphasis on the dorsal horn.

  Authors: A J Todd, D I Hughes, E Polgár, G G Nagy, M Mackie, O P Ottersen, D J Maxwell

  The European journal of neuroscience. 02/2003; 17(1):13-27.

  Two vesicular glutamate transporters, VGLUT1 and VGLUT2, have recently been identified, and it has been reported that they are expressed by largely nonoverlapping populations of glutamatergic neuronsTwo vesicular glutamate transporters, VGLUT1 and VGLUT2, have recently been identified, and it has been reported that they are expressed by largely nonoverlapping populations of glutamatergic neurons in the brain. We have used immunocytochemistry with antibodies against both transporters, together with markers for various populations of spinal neurons, in an attempt to identify glutamatergic interneurons in the dorsal horn of the mid-lumbar spinal cord of the rat. The great majority (94-100%) of nonprimary axonal boutons that contained somatostatin, substance P or neurotensin, as well as 85% of those that contained enkephalin, were VGLUT2-immunoreactive, which suggests that most dorsal horn neurons that synthesize these peptides are glutamatergic. In support of this, we found that most somatostatin- and enkephalin-containing boutons (including somatostatin-immunoreactive boutons that lacked calcitonin gene-related peptide and were therefore probably derived from local interneurons) formed synapses at which AMPA receptors were present. We also investigated VGLUT expression in central terminals of primary afferents. Myelinated afferents were identified with cholera toxin B subunit; most of those in lamina I were VGLUT2-immunoreactive, whereas all those in deeper laminae were VGLUT1-immunoreactive, and some (in laminae III-VI) appeared to contain both transporters. However, peptidergic primary afferents that contained substance P or somatostatin (most of which are unmyelinated), as well as nonpeptidergic C fibres (identified with Bandeiraea simplicifolia isolectin B4) showed low levels of VGLUT2-immunoreactivity, or were not immunoreactive with either VGLUT antibody. As all primary afferents are thought to be glutamatergic, this raises the possibility that unmyelinated afferents, most of which are nociceptors, express a different vesicular glutamate transporter.

• Myelinated and unmyelinated primary afferent axons form contacts with cholinergic interneurons in the spinal dorsal horn.

  Authors: M J Olave, N Puri, R Kerr, D J Maxwell

  Experimental brain research. Experimentelle Hirnforschung. Expérimentation cérébrale. 09/2002; 145(4):448-56.

  Cholinergic interneurons in laminae III/IV of the dorsal horn contain co-localised gamma-aminobutyric acid (GABA) and frequently form axoaxonic synapses with terminals of primary afferents. They areCholinergic interneurons in laminae III/IV of the dorsal horn contain co-localised gamma-aminobutyric acid (GABA) and frequently form axoaxonic synapses with terminals of primary afferents. They are therefore probably last-order interneurons involved in presynaptic inhibition. The purpose of the present investigation was to determine if these cells receive direct input from primary afferents. Relationships between primary afferents and interneurons were investigated in adult rats. Myelinated primary afferents were labelled with the B-subunit of cholera toxin (CTb). Unmyelinated afferents were labelled with IB4 lectin and an antibody to identify calcitonin-gene-related peptide (CGRP). Cholinergic neurons were labelled with an antibody raised against choline acetyltransferase and examined with a confocal microscope. Cells were reconstructed with NeuroLucida for Confocal and afferent contacts plotted. Interneurons (N=30) received an average of 20.2+/-11.9 (SD) contacts from CTb-labelled primary afferents, which were preferentially distributed on proximal and intermediate dendrites. Interneurons with dendrites which extended into lamina II (N=20) received an average of 27.4+/-19.0 IB4 contacts (on intermediate and distal dendrites) and 9.2+/-6.8 CGRP contacts. It is concluded that cholinergic interneurons receive contacts from both myelinated and unmyelinated primary afferents and different classes of afferent target particular dendritic domains. Cholinergic interneurons are likely to be components of an inhibitory feedback pathway that is monosynaptically activated by primary afferents.

• An investigation of neurones that possess the alpha 2C-adrenergic receptor in the rat dorsal horn.

  Authors: M J Olave, D J Maxwell

  Neuroscience. 02/2002; 115(1):31-40.

  The function of the alpha(2C) subclass of adrenergic receptor in the spinal cord is unclear at present. Immunoreactivity for this receptor is found predominantly on axon terminals of the superficialThe function of the alpha(2C) subclass of adrenergic receptor in the spinal cord is unclear at present. Immunoreactivity for this receptor is found predominantly on axon terminals of the superficial dorsal horn but limited information is available about the properties and origin of these axons. The aim of this study was to determine which classes of neurone give rise to axons that possess this receptor and to investigate the synaptic organisation of these terminals. A series of double-labelling experiments was performed to investigate the relationship between the alpha(2C) receptor and each one of 14 chemical markers that label various types of axon terminal in the dorsal horn. Tissue was examined with two-colour confocal laser scanning microscopy. Quantitative analysis revealed that alpha(2C)-adrenergic receptors are not present on terminals of unmyelinated or peptidergic primary afferents and descending noradrenergic or serotoninergic axons. They were found on a proportion of terminals belonging to a mixed population of excitatory and inhibitory spinal interneurones, including those that contain neurotensin, somatostatin, enkephalin, GABA and neuropeptide Y. However, a greater proportion of terminals originating from excitatory interneurones were found to possess the receptor. Electron microscopic analysis revealed that alpha(2C)-adrenergic receptor immunoreactivity is predominantly associated with axon terminals that are presynaptic to dendrites but a small proportion of immunoreactive terminals formed axo-axonic synaptic arrangements.These studies indicate that noradrenaline can modulate transmission in the dorsal horn by acting through alpha(2C)-adrenergic receptors on terminals of spinal interneurones.

• Morphological evidence for selective modulation by serotonin of a subpopulation of dorsal horn cells which possess the neurokinin-1 receptor.

  Authors: W Stewart, D J Maxwell

  The European journal of neuroscience. 01/2001; 12(12):4583-8.

  Serotonin selectively depresses transmission of nociceptive information through the spinal dorsal horn but the mechanisms of this depression are poorly understood. In this study we report thatSerotonin selectively depresses transmission of nociceptive information through the spinal dorsal horn but the mechanisms of this depression are poorly understood. In this study we report that serotonin-containing axons form basket-like clusters which are intimately woven around cell bodies and proximal dendrites of a subpopulation ( approximately 50%) of laminae III/IV neurons which possess the neurokinin-1 receptor. Statistical analysis confirms that cells belonging to this subpopulation have significantly higher numbers of serotoninergic contacts on proximal dendrites when compared with the population of neurokinin-1 cells that are not associated with clusters (mean +/- SD = 13+/-5.8 and 5+/-2.9, respectively). Neurokinin-1 cells in laminae III/IV project to regions of the brain which are involved in nociceptive processing and are likely to be activated predominantly by nociceptive input. The concentration of serotoninergic axons around proximal regions of some of these cells indicates that serotonin may have a powerful influence on transmission through this pathway. This type of arrangement could be a morphological correlate for at least part of the selective antinociceptive actions of serotonin.

• Coupling between serotoninergic and noradrenergic neurones and gamma-motoneurones in the cat.

  Authors: M H Gladden, D J Maxwell, A Sahal, E Jankowska

  The Journal of physiology. 10/2000; 527 Pt 2:213-23.

  Noradrenaline is known to suppress transmission from group II muscle afferents when locally applied to gamma-motoneurones, and serotonin (5-HT) facilitates the transmission. The purpose of thisNoradrenaline is known to suppress transmission from group II muscle afferents when locally applied to gamma-motoneurones, and serotonin (5-HT) facilitates the transmission. The purpose of this investigation was to search for evidence of monoaminergic innervation of gamma-motoneurones. Eight gamma-motoneurones were labelled with rhodamine-dextran, and 50 micrometer thick sagittal sections of the spinal cord containing them were exposed to antibodies against dopamine beta-hydroxylase (DBH) and 5-HT. All the cells were directly and/or indirectly excited by muscle group II afferents from the muscle they innervated and/or other muscles. Appositions between monoaminergic fibres and the labelled somata and dendrites were located with three-colour confocal laser scanning microscopy by examining series of optical sections at 1 or 0.5 micrometer intervals. DBH and 5-HT varicosities formed appositions with the somata and dendrites of all the gamma-motoneurones. The mean packing densities for 5-HT (1.12 +/- 0.11 appositions per 100 micrometer(2) for somata and 0.91 +/- 0.07 per 100 micrometer(2) for dendrites) were similar to the densities of contacts reported for alpha-motoneurones. Monoaminergic varicosities in apposition to dendrites greatly outnumbered those on the somata. The density of DBH appositions was consistently lower - corresponding means were 53% and 62% of those for 5-HT on the somata and dendrites, respectively. It is concluded from an analysis of the distribution and density of varicosities in apposition to the gamma-motoneurones compared with the density in the immediate surround of the dendrites that there is indeed both a serotoninergic and noradrenergic innervation of gamma-motoneurones.

• Serotoninergic and noradrenergic axonal contacts associated with premotor interneurons in spinal pathways from group II muscle afferents.

  Authors: D J Maxwell, J S Riddell, E Jankowska

  The European journal of neuroscience. 05/2000; 12(4):1271-80.

  We investigated the possibility that monoaminergic axons make contacts with spinal interneurons which project to motor nuclei and are monosynaptically activated by group II muscle afferents.We investigated the possibility that monoaminergic axons make contacts with spinal interneurons which project to motor nuclei and are monosynaptically activated by group II muscle afferents. Interneurons in midlumbar spinal segments of adult cats were characterized electrophysiologically and intracellularly labelled with tetramethylrhodamine dextran. Serotoninergic and noradrenergic axons were identified with immunofluorescence in sections containing labelled cells. Contacts between monoaminergic axons and interneurons were investigated with three-colour confocal laser scanning microscopy and analysed with a computer reconstruction program. Cell bodies and dendritic trees of five cells were reconstructed and putative contacts were plotted. The average number of contacts formed by serotoninergic axons was 140 and the average number of noradrenergic contacts was 38. The majority (95%) of contacts were formed with dendrites; these were distributed over the entire dendritic tree, even on the most distal branches. These findings provide a morphological basis for the modulatory actions of monoamines on premotor spinal interneurons in pathways from group II muscle afferents.

• Axoaxonic synapses on terminals of group II muscle spindle afferent axons in the spinal cord of the cat.

  Authors: D J Maxwell, J S Riddell

  The European journal of neuroscience. 07/1999; 11(6):2151-9.

  The purpose of the present study was to determine if terminals of identified group II muscle spindle afferents participate in axoaxonic synaptic arrangements and, if so, to investigate theThe purpose of the present study was to determine if terminals of identified group II muscle spindle afferents participate in axoaxonic synaptic arrangements and, if so, to investigate the transmitter content of presynaptic terminals in these arrangements. Group II muscle afferents supplying the gastrocnemius-soleus or semitendinosus muscles were identified in adult cats and stained intra-axonally with horseradish peroxidase. In total, three group II axons were labelled and processed for combined light and electron microscopy. Group II axons gave rise to collaterals which characteristically descended through the superficial dorsal horn and formed relatively sparse terminal arborizations in the dorsal horn (laminae IV and V) and more profuse arbors in the intermediate grey matter (laminae VI-VII). Forty boutons were examined through series of ultrathin sections and all but four were postsynaptic to other axon terminals. Occasionally, more than one axon was presynaptic to a single group II terminal. Immunogold studies showed that all axons in presynaptic apposition to group II boutons contained gamma-aminobutyric acid (GABA) and also that glycine was colocalized in the majority of these axons. This evidence suggests that transmission from group II muscle afferents is under strong presynaptic inhibitory control and that it is mainly the subgroup of GABAergic interneurons with colocalized glycine which mediate this inhibition. Seventeen group II boutons were components of synaptic triads where the presynaptic axoaxonic bouton formed a synapse with the same dendrite as the group II axon. Therefore, a proportion of the interneurons which form axoaxonic synapses with group II axons are also likely to have postsynaptic inhibitory actions on target neurons of group II afferents.

• Differential expression of the muscarinic m2 acetylcholine receptor by small and large motoneurons of the rat spinal cord.

  Authors: J Welton, W Stewart, R Kerr, D J Maxwell

  Brain research. 02/1999; 817(1-2):215-9.

  The purpose of this study was to determine if motoneurons in the spinal ventral horn express the muscarinic m2 acetylcholine receptor. Motoneurons were retrogradely labelled in adult rats byThe purpose of this study was to determine if motoneurons in the spinal ventral horn express the muscarinic m2 acetylcholine receptor. Motoneurons were retrogradely labelled in adult rats by intramuscular injection of Fluorogold and examined with confocal microscopy for evidence of immunoreactivity for the receptor. The cells were also double-labelled for choline acetyltransferase to determine if they were contacted by cholinergic axons. Almost all large motoneurons (diameters greater than 35 microm) displayed intense immunoreactivity for the receptor which was evenly distributed along the plasma membrane. Small cells were immunonegative for the receptor or weakly labelled. As large cells probably correspond to alpha-motoneurons and receive many cholinergic contacts, it is concluded that acetylcholine can influence this type of motoneuron by acting through the m2 receptor.