Neuronal synchrony in the basolateral amygdala (BLA) is critical for emotional behavior. Coordinated theta-frequency oscillations between the BLA and the hippocampus and precisely timed integration of salient sensory stimuli in the BLA are involved in fear conditioning. We characterized GABAergic interneuron types of the BLA and determined their contribution to shaping these network activities. Using in vivo recordings in rats combined with the anatomical identification of neurons, we found that the firing of BLA interneurons associated with network activities was cell type specific. The firing of calbindin-positive interneurons targeting dendrites was precisely theta-modulated, but other cell types were heterogeneously modulated, including parvalbumin-positive basket cells. Salient sensory stimuli selectively triggered axo-axonic cells firing and inhibited firing of a disctinct projecting interneuron type. Thus, GABA is released onto BLA principal neurons in a time-, domain-, and sensory-specific manner. These specific synaptic actions likely cooperate to promote amygdalo-hippocampal synchrony involved in emotional memory formation.
... Interestingly, superficial layer 2/3 (L2/3) PyCs in V1 also receive more ChC synapses than deep layer 3 PyCs (13), but whether connections are non-reciprocal remains unknown. Recent studies using in vivo twophoton calcium imaging in V1 show that ChC activity is highly correlated with pupil size and locomotion (13,30), indicating arousal-related ChC activity consistent with what has been observed in other brain regions (28,31,32). ...
... A third option is that the experience-dependent changes in ChC activity are related to arousal. ChCs were previously shown to display arousal-related activity in various brain regions (13,28,(30)(31)(32), which matches our observation that their activity is highly correlated with running and pupil size. In addition, the neuromodulator acetylcholine (ACh) is involved in axonal arborization of developing ChCs, further underlining the relationship between ChCs and arousal (68). ...
Detailed characterization of interneuron types in primary visual cortex (V1) has greatly contributed to understanding visual perception, yet the role of chandelier cells (ChCs) in visual processing remains poorly characterized. Using viral tracing we found that V1 ChCs predominantly receive monosynaptic input from local layer 5 pyramidal cells and higher-order cortical regions. Two-photon calcium imaging and convolutional neural network modelling revealed that ChCs are visually responsive but weakly selective for stimulus content. In mice running in a virtual tunnel, ChCs respond strongly to events known to elicit arousal, including locomotion and visuomotor mismatch. Repeated exposure of the mice to the virtual tunnel was accompanied by reduced visual responses of ChCs and structural plasticity of ChC boutons and axon initial segment length. Finally, ChCs only weakly inhibited pyramidal cells. These findings suggest that ChCs provide an arousal-related signal to layer 2/3 pyramidal cells that may modulate their activity and/or gate plasticity of their axon initial segments during behaviorally relevant events.
... This defining feature of synapse specificity led to subsequent discoveries of AACs in the hippocampus and BLA. These findings have relied on meticulous efforts of skilled neuroanatomists to correctly identify AACs, mostly post hoc, by synapse specificity at AIS (Bienvenu et al., 2012 ;Veres et al., 2014 ). Since then, it has been widely assumed that AACs are restricted to these brain structures, and likely to the mammalian species. ...
Axo-axonic cells (AACs), also called chandelier cells (ChCs) in the cerebral cortex, are the most distinctive type of GABAergic interneurons described in the neocortex, hippocampus, and basolateral amygdala (BLA). AACs selectively innervate glutamatergic projection neurons (PNs) at their axon initial segment (AIS), thus may exert decisive control over PN spiking and regulate PN functional ensembles. However, the brain-wide distribution, synaptic connectivity, and circuit function of AACs remains poorly understood, largely due to the lack of specific and reliable experimental tools. Here, we have established an intersectional genetic strategy that achieves specific and comprehensive targeting of AACs throughout the mouse brain based on their lineage ( Nkx2.1 ) and molecular ( Unc5b , Pthlh ) markers. We discovered that AACs are deployed across essentially all the pallium-derived brain structures, including not only the dorsal pallium-derived neocortex and medial pallium-derived hippocampal formation, but also the lateral pallium-derived claustrum-insular complex, and the ventral pallium-derived extended amygdaloid complex and olfactory centers. AACs are also abundant in anterior olfactory nucleus, taenia tecta and lateral septum. AACs show characteristic variations in density across neocortical areas and layers and across subregions of the hippocampal formation. Neocortical AACs comprise multiple laminar subtypes with distinct dendritic and axonal arborization patterns. Retrograde monosynaptic tracing from AACs across neocortical, hippocampal and BLA regions reveal shared as well as distinct patterns of synaptic input. Specific and comprehensive targeting of AACs facilitates the study of their developmental genetic program and circuit function across brain structures, providing a ground truth platform for understanding the conservation and variation of a bona fide cell type across brain regions and species.
... Several studies indicate that different afferents to the BLA target discrete projecting populations that consequently modulate distinct behaviors. 88 While we did not explore interneuron diversity 103,104 in regard to the different cortical inputs in this work, recent studies have highlighted key roles for inhibitory neuron subtypes in information processing and disinhibition in the BLA during various aspects of anxiety and fear. 102,[105][106][107][108][109][110] Our data also suggests a role for various forms of synaptic plasticity in underlying the behavioral response to mGluR2 activation. ...
Dissecting how membrane receptors regulate neural circuit function is critical for deciphering basic principles of neuromodulation and mechanisms of therapeutic drug action. Classical pharmacological and genetic approaches are not well-equipped to untangle the roles of specific receptor populations, especially in long-range projections which coordinate communication between brain regions. Here we use viral tracing, electrophysiological, optogenetic, and photopharmacological approaches to determine how presynaptic metabotropic glutamate receptor 2 (mGluR2) activation in the basolateral amygdala (BLA) alters anxiety-related behavior. We find that mGluR2-expressing neurons from the ventromedial prefrontal cortex (vmPFC) and posterior insular cortex (pIC) preferentially target distinct cell types and subregions of the BLA to regulate different forms of avoidant behavior. Using projection-specific photopharmacological activation, we find that mGluR2-mediated presynaptic inhibition of vmPFC-BLA, but not pIC-BLA, connections can produce long-lasting decreases in spatial avoidance. In contrast, presynaptic inhibition of pIC-BLA connections decreased social avoidance, novelty-induced hypophagia, and increased exploratory behavior without impairing working memory, establishing this projection as a novel target for the treatment of anxiety disorders. Overall, this work reveals new aspects of BLA neuromodulation with therapeutic implications while establishing a powerful approach for optical mapping of drug action via photopharmacology.
Highlights
- Basolateral amygdala is a key site for anxiolytic action of mGluR2 agonism
- BLA receives dense Grm2 ⁺ inputs from ventromedial prefrontal cortex and posterior insular cortex
- Photoactivation of mGluR2 has distinct anxiolytic effects in vmPFC-BLA and pIC-BLA synapses
- Grm2 ⁺ vmPFC and pIC projections target medial and lateral BLA subregions, respectively
Analysis of cytoarchitectonics of the basolateral nucleus (BLN) of the brain amygdala was performed in cresyl violet-stained frontal paraffin sections of the brain in 10 alcohol-preferring (AP) and 10 alcohol-nonpreferring (ANP) rats (with an equal number of male and female animals in each group). The presence of large and small neurons was detected in BLN. Most of the large neurons in AP rats had the character of chromoneutral and moderately chromophilic cells, while in ANP rats these cells were moderately chromophobic. Application of Golgi method demonstrated that the equivalents of large neurons were long-axonal densely branched pyramid-like neurons, and those of small-sized neurons - short-axonal neurons. The determination of the ratio of large and small-sized neurons showed that in AP rats the proportion of latter was 12.3±0.6%, while in the ANP rats it was significantly greater - 19.70±0.23%. These results help to explain the previously obtained data on larger specific area of BLN in amygdala of ANP rats by the presence of greater number of interneurons than in AP rats.
Acetylcholine (ACh) is released from basal forebrain cholinergic neurons in response to salient stimuli and engages brain states supporting attention and memory. These high ACh states are associated with theta oscillations, which synchronize neuronal ensembles. Theta oscillations in the basolateral amygdala (BLA) in both humans and rodents have been shown to underlie emotional memory, yet their mechanism remains unclear. Here, using brain slice electrophysiology in male and female mice, we show large ACh stimuli evoke prolonged theta oscillations in BLA local field potentials that depend upon M3 muscarinic receptor activation of cholecystokinin (CCK) interneurons (INs) without the need for external glutamate signaling. Somatostatin (SOM) INs inhibit CCK INs and are themselves inhibited by ACh, providing a functional SOM→CCK IN circuit connection gating BLA theta. Parvalbumin (PV) INs, which can drive BLA oscillations in baseline states, are not involved in the generation of ACh-induced theta, highlighting that ACh induces a cellular switch in the control of BLA oscillatory activity and establishes an internally BLA-driven theta oscillation through CCK INs. Theta activity is more readily evoked in BLA over the cortex or hippocampus, suggesting preferential activation of the BLA during high ACh states. These data reveal a SOM→CCK IN circuit in the BLA that gates internal theta oscillations and suggest a mechanism by which salient stimuli acting through ACh switch the BLA into a network state enabling emotional memory.
Basket cells are inhibitory interneurons in cortical structures with the potential to efficiently control the activity of their postsynaptic partners. Although their contribution to higher order cognitive functions associated with the medial prefrontal cortex (mPFC) relies on the characteristics of their synaptic connections, the way they are embedded into local circuits is still not fully uncovered. Here, we determined the synaptic properties of excitatory and inhibitory connections between pyramidal neurons (PNs), cholecystokinin-containing basket cells (CCKBCs) and parvalbumin-containing basket cells (PVBCs) in the mouse mPFC. By performing paired recordings, we revealed that PVBCs receive larger unitary excitatory postsynaptic currents from PNs with shorter latency and faster kinetic properties compared to events evoked in CCKBCs. Also, unitary inhibitory postsynaptic currents in PNs were more reliably evoked by PVBCs than by CCKBCs yet the former connections showed profound short-term depression. Moreover, we demonstrated that CCKBCs and PVBCs in the mPFC are mutually interconnected with each other. As alterations in PVBC function have been linked to neurological and psychiatric conditions like Alzheimer disease and schizophrenia and CCKBC vulnerability might play a role in mood disorders, a deeper understanding of the general features of basket cell synapses could serve as a reference point for future investigations with therapeutic objectives.
Detailed characterization of interneuron types in primary visual cortex (V1) has greatly contributed to understanding visual perception, yet the role of chandelier cells (ChCs) in visual processing remains poorly characterized. Using viral tracing we found that V1 ChCs predominantly receive monosynaptic input from local layer 5 pyramidal cells and higher-order cortical regions. Two-photon calcium imaging and convolutional neural network modeling revealed that ChCs are visually responsive but weakly selective for stimulus content. In mice running in a virtual tunnel, ChCs respond strongly to events known to elicit arousal, including locomotion and visuomotor mismatch. Repeated exposure of the mice to the virtual tunnel was accompanied by reduced visual responses of ChCs and structural plasticity of ChC boutons and axon initial segment length. Finally, ChCs only weakly inhibited pyramidal cells. These findings suggest that ChCs provide an arousal-related signal to layer 2/3 pyramidal cells that may modulate their activity and/or gate plasticity of their axon initial segments during behaviorally relevant events.
Memory formation depends on neural activity across a network of regions, including the hippocampus and broader medial temporal lobe (MTL). Interactions between these regions have been studied indirectly using functional MRI, but the bases for interregional communication at a cellular level remain poorly understood. Here we evaluate the hypothesis that oscillatory currents in the hippocampus synchronize the firing of neurons both within and outside the hippocampus. We recorded extracellular spikes from 1,854 single- and multi-units simultaneously with hippocampal local field potentials (LFPs) in 28 neurosurgical patients who completed virtual navigation experiments. A majority of hippocampal neurons phase-locked to oscillations in the slow (2-4Hz) or fast (6-10Hz) theta bands, with a significant subset exhibiting nested slow theta x beta frequency (13-20Hz) phase-locking. Outside of the hippocampus, phase-locking to hippocampal oscillations occurred only at theta frequencies and primarily among neurons in the entorhinal cortex and amygdala. Moreover, extrahippocampal neurons phase-locked to hippocampal theta even when theta did not appear locally. These results indicate that spike-time synchronization with hippocampal theta is a defining feature of neuronal activity in the hippocampus and structurally connected MTL regions. Theta phase-locking could mediate flexible communication with the hippocampus to influence the content and quality of memories.
Axo-axonic cells (AACs), also called chandelier cells (ChCs) in the cerebral cortex, are the most distinctive type of GABAergic interneurons described in the neocortex, hippocampus, and basolateral amygdala (BLA). AACs selectively innervate glutamatergic projection neurons (PNs) at their axon initial segment (AIS), thus may exert decisive control over PN spiking and regulate PN functional ensembles. However, the brain-wide distribution, synaptic connectivity, and circuit function of AACs remains poorly understood, largely due to the lack of specific and reliable experimental tools. Here, we have established an intersectional genetic strategy that achieves specific and comprehensive targeting of AACs throughout the mouse brain based on their lineage ( Nkx2.1 ) and molecular ( Unc5b , Pthlh ) markers. We discovered that AACs are deployed across essentially all the pallium-derived brain structures, including not only the dorsal pallium-derived neocortex and medial pallium-derived hippocampal formation, but also the lateral pallium-derived claustrum-insular complex, and the ventral pallium-derived extended amygdaloid complex and olfactory centers. AACs are also abundant in anterior olfactory nucleus, taenia tecta and lateral septum. AACs show characteristic variations in density across neocortical areas and layers and across subregions of the hippocampal formation. Neocortical AACs comprise multiple laminar subtypes with distinct dendritic and axonal arborization patterns. Retrograde monosynaptic tracing from AACs across neocortical, hippocampal and BLA regions reveal shared as well as distinct patterns of synaptic input. Specific and comprehensive targeting of AACs facilitates the study of their developmental genetic program and circuit function across brain structures, providing a ground truth platform for understanding the conservation and variation of a bona fide cell type across brain regions and species.
Detailed characterization of interneuron subtypes in primary visual cortex (V1) has greatly contributed to understanding visual perception, yet the role of chandelier cells (ChCs) in visual processing remains poorly characterized. Using viral tracing we found that V1 ChCs predominantly receive monosynaptic input from local layer 5 pyramidal cells and higher-order cortical regions. Two-photon calcium imaging and convolutional neural network modelling revealed that ChCs are visually responsive but weakly selective for stimulus content. In mice running in a virtual tunnel, ChCs respond strongly to locomotion and halting visual flow, suggesting arousal-related activity. Visuomotor experience in the tunnel diminished visual responses of ChCs and induced structural plasticity of ChC boutons and axon initial segment length. Finally, ChCs only weakly inhibited pyramidal cells. These findings suggest that ChCs provide an arousal-related signal to layer 2/3 pyramidal cells that may modulate their activity and/or gate plasticity of their axon initial segments during behaviorally relevant events.
The hippocampus (CA1, CA2, and CA3 areas and the dentate gyrus), together with the subiculum, represents an associational area of the cerebral cortex intimately involved in mnemonic processes. Through its connections with other areas of the temporal lobe, the hippocampus contributes to the encoding, association, consolidation, and recall of representations of the external and internal world in the combined firing rates and spike timing of glutamatergic pyramidal and granule cells. The hippocampus is thought to associate specific life events (items, episodes), on several time scales, in temporally determined firing sequences of neuronal assemblies. A single pyramidal cell can be part of several cell assemblies with different partners and contribute to different representations. Pyramidal cell assemblies are thought to be kept together and segregated from other assemblies by the dynamic strengthening and weakening of glutamatergic synaptic weights as well as by GABAergic interneurons. Interneurons generate cell domain and brain state-dependent rhythmic changes in excitability, which are key for the formation, consolidation, and recall of representations. Unsurprisingly, interneurons show intricate spatiotemporal diversity; for example, the CA1 area is served by at least twenty-one types of resident GABAergic cell. This chapter attempts to allocate explicit roles for some of them, based on their previously published firing patterns in vivo as observed in identified neurons recorded in anesthetized rats and on their putative equivalents in nonanesthetized animals.
Neural-network oscillations at distinct frequencies have been implicated in the encoding, consolidation and retrieval of information in the hippocampus. Some GABA (γ-aminobutyric acid)-containing interneurons fire phase-locked to theta oscillations (4-8Hz) or to sharp-wave-associated ripple oscillations (120-200 Hz), which represent different behavioural states. Interneurons also entrain pyramidal cells in vitro. The large diversity of interneurons poses the question of whether they have specific roles in shaping distinct network activities in vivo. Here we report that three distinct interneuron types - basket, axo-axonic and oriens-lacunosum-moleculare cells - visualized and defined by synaptic connectivity as well as by neurochemical markers, contribute differentially to theta and ripple oscillations in anaesthetized rats. The firing patterns of individual cells of the same class are remarkably stereotyped and provide unique signatures for each class. We conclude that the diversity of interneurons, innervating distinct domains of pyramidal cells, emerged to coordinate the activity of pyramidal cells in a temporally distinct and brain-state-dependent manner.
Immunohistochemical double-label techniques were used to study the localization of DARPP-32, a phosphoprotein that is enriched in neurons possessing members of the D1 subfamily of dopamine receptors, in several different types of striatal neurons in the rat basal ganglia. The vast majority (94.1%) of striatonigral projection neurons (the vast majority of which contain substance P), identified by retrograde labeling with fluorogold, were observed to contain DARPP-32. Similarly, the vast majority of striatopallidal projection neurons (87.7%), identified by immunofluorescence labeling for enkephalin (ENK), were found to label for DARPP-32. In contrast, cholinergic and neuropeptide Y-containing striatal interneurons were never observed to contain DARPP-32. These results suggest that essentially all major types of striatal medium spiny projection neurons may possess members of the D1 subfamily of dopamine receptors, but that striatal local circuit neurons do not possess members of the D1 subfamily of receptors.
Transforming synaptic input into action potential output is a fundamental function of neurons. The pattern of action potential output from principal cells of the mammalian hippocampus encodes spatial and nonspatial information, but the cellular and circuit mechanisms by which neurons transform their synaptic input into a given output are unknown. Using a combination of optical activation and cell type-specific pharmacogenetic silencing in vitro, we found that dendritic inhibition is the primary regulator of input-output transformations in mouse hippocampal CA1 pyramidal cells, and acts by gating the dendritic electrogenesis driving burst spiking. Dendrite-targeting interneurons are themselves modulated by interneurons targeting pyramidal cell somata, providing a synaptic substrate for tuning pyramidal cell output through interactions in the local inhibitory network. These results provide evidence for a division of labor in cortical circuits, where distinct computational functions are implemented by subtypes of local inhibitory neurons.
Chandelier (axoaxonic) cells (ChCs) are a distinct group of GABAergic interneurons that innervate the axon initial segments of pyramidal cells. However, their circuit role and the function of their clearly defined anatomical specificity remain unclear. Recent work has demonstrated that chandelier cells can produce depolarizing GABAergic PSPs, occasionally driving postsynaptic targets to spike. On the other hand, other work suggests that ChCs are hyperpolarizing and may have an inhibitory role. These disparate functional effects may reflect heterogeneity among ChCs. Here, using brain slices from transgenic mouse strains, we first demonstrate that, across different neocortical areas and genetic backgrounds, upper Layer 2/3 ChCs belong to a single electrophysiologically and morphologically defined population, extensively sampling Layer 1 inputs with asymmetric dendrites. Consistent with being a single cell type, we find electrical coupling between ChCs. We then investigate the effect of chandelier cell activation on pyramidal neuron spiking in several conditions, ranging from the resting membrane potential to stimuli designed to approximate in vivo membrane potential dynamics. We find that under quiescent conditions, chandelier cells are capable of both promoting and inhibiting spike generation, depending on the postsynaptic membrane potential. However, during in vivo-like membrane potential fluctuations, the dominant postsynaptic effect was a strong inhibition. Thus, neocortical chandelier cells, even from within a homogeneous population, appear to play a dual role in the circuit, helping to activate quiescent pyramidal neurons, while at the same time inhibiting active ones.
Neurons in the lateral and basolateral nuclei of the rat amygdala were studied using Golgi-Kopsch and rapid Golgi techniques. According to differences in perikaryal, dendritic, and axonal morphology, three main neuronal classes are recognized. Class I neurons, the predominant cell type in both nuclei, are large, spiny neurons that vary in size in different subdivisions of the lateral and basolateral nuclei. These neurons often have a pyramidal shape, exhibiting one or two thick “apical” dendrites and several thinner “basal” dendrites. Axons of class I neurons, which appear to pass out of the nucleus of origin, usually give off several collaterals that arborize modestly in the vicinity of the cell. Class II neurons are smaller, ovoid cells that comprise approximately 5% of impregnated neurons. These neurons are characterized by spine-sparse dendrites and fairly dense local axonal arborizations. Class II neurons may be classified as multipolar, bitufted, or bipolar, depending on dendritic branching pattern. Another type of class II neuron, the amygdaloid chandelier cell, is recognized by virtue of its distinctive axon. The chandelier cell axon gives off numerous collaterals that form nestlike entanglements exhibiting clusters of axonal varicosities. Isolated chandelierlike axons of undetermined origin were observed forming multiple contacts with initial segments of class I axons. Several small, spherical class III neurons with short, varicose dendrites were observed. Axons branch profusely to form a dense tangle of collaterals in the vicinity of the cell. Both axons and dendrites establish numerous contacts with class I dendrites. This investigation, the first detailed Golgi study of the basolateral amygdala of the rat, reveals that the cytoarchitecture of this brain region in the rat is basically similar to that of the opossum and other mammals. Morphologic details described in this report should be useful in the interpretation of ultrastructural, immunocytochemical, and electro-physiological studies of the basolateral amygdala.
