Pyramidal Cell Somata Research Articles

Responses of pyramidal cell somata and apical dendrites in mouse visual cortex over multiple days

The apical dendrites of pyramidal neurons in sensory cortex receive primarily top-down signals from associative and motor regions, while cell bodies and nearby dendrites are heavily targeted by locally recurrent or bottom-up inputs from the sensory periphery. Based on these differences, a number of theories in computational neuroscience postulate a unique role for apical dendrites in learning. However, due to technical challenges in data collection, little data is available for comparing the responses of apical dendrites to cell bodies over multiple days. Here we present a dataset collected through the Allen Institute Mindscope’s OpenScope program that addresses this need. This dataset comprises high-quality two-photon calcium imaging from the apical dendrites and the cell bodies of visual cortical pyramidal neurons, acquired over multiple days in awake, behaving mice that were presented with visual stimuli. Many of the cell bodies and dendrite segments were tracked over days, enabling analyses of how their responses change over time. This dataset allows neuroscientists to explore the differences between apical and somatic processing and plasticity.

Immunohistochemical Identification of Interneuronal Subpopulations in the Basolateral Amygdala of the Rhesus Monkey (Macaca mulatta)

Inhibitory circuits in the basolateral nuclear complex of the amygdala (BNC) critical for controlling the acquisition, expression, and extinction of emotional responses are mediated by GABAergic interneurons (INs). Studies in rodents have demonstrated that separate IN subpopulations, identified by their expression of calcium-binding proteins and neuropeptides, play discrete roles in the intrinsic circuitry of the BNC. Far less is known about IN subpopulations in primates. In order to fill in this gap in our understanding of primate INs, the present investigation used dual-labeling immunohistochemistry for IN markers to identify subpopulations expressing cholecystokinin (CCK), calbindin (CB), calretinin (CR), and somatostatin (SOM) in somata and axon terminals in the monkey BNC. In general, colocalization patterns seen in somata and axon terminals were similar. It was found that there was virtually no colocalization of CB and CR, the two calcium-binding proteins investigated. Three subtypes of CCK-immunoreactive (CCK+) INs were identified on the basis of their expression of CR or CB: (1) CCK+/CR+; (2) CCK+/CB+); and (3) CCK+/CR−/CB−. Almost no colocalization of CCK with SOM was observed, but there was extensive colocalization of SOM and CB. CCK+, CR+, and CCK+/CR+ double-labeled axon terminals were seen surrounding pyramidal cell somata in basket-like plexuses, as well as in the neuropil. CB+, SOM+, and CB+/SOM+ terminals did not form baskets, suggesting that these IN subpopulations are mainly dendrite-targeting neurons. In general, the IN subpopulations in the monkey are not dissimilar to those seen in rodents but, unlike rodents, CB+ INs in the monkey are not basket cells.

Cholecystokinin immunoreactive neurons in the basolateral amygdala of the rhesus monkey (Macaca mulatta).

Several distinct subpopulations of interneurons (INs) in the amygdalar basolateral nuclear complex (BNC) of the rat can be recognized on the basis of their expression of calcium-binding proteins and neuropeptides, including parvalbumin (PV), somatostatin (SOM), calretinin (CR), and cholecystokinin (CCK). In the rat BNC CCK is expressed in two separate IN subpopulations, termed large (CCKL ) and small (CCKS ). These subpopulations exhibit distinct connections indicative of discrete functional roles in the circuitry of the BNC. Although there have been several studies of PV+, SOM+, and CR+ INs in the primate BNC, there is almost no information regarding CCK+ INs in these species. Therefore, in the present study the distribution and morphology of CCK+ INs and their axon terminals in the BNC of the monkey was investigated. CCK immunoreactivity in the BNC was observed in somata and proximal dendrites of nonpyramidal neurons, as well as in axon terminals. A moderate density of CCK+ INs was found in all nuclei of the BNC. CCK+ INs in the BNC were morphologically heterogeneous, with both small and large varieties observed. All CCK+ somata gave rise to 2-4 dendrites that branched sparingly and were aspiny. CCK+ axon terminals in the BNC were found both in the neuropil and forming pericellular baskets contacting somata of pyramidal cells. In addition, many CCK+ neurons were contacted by multiple CCK+ terminals, indicative of the existence of a CCK interneuronal network. These data indicate that the morphology of CCK+ INs in the monkey is very similar to that of the rat.

TRPC1 Channels Are Expressed in Pyramidal Neurons and in a Subset of Somatostatin Interneurons in the Rat Neocortex

Disturbances in calcium homeostasis due to canonical transient receptor potential (TRPC) and/or store-operated calcium (SOC) channels can play a key role in a large number of brain disorders. TRPC channels are plasma membrane cation channels included in the transient receptor potential (TRP) superfamily. The most widely distributed member of the TRPC subfamily in the brain is TRPC1, which is frequently linked to group I metabotropic glutamate receptors (mGluRs) and to the components of SOC channels. Proposing TRPC/SOC channels as a therapeutic target in neurological diseases previously requires a detailed knowledge of the distribution of such molecules in the brain. The aim of our study was to analyze the neuroanatomical distribution of TRPC1 in the rat neocortex. By double- and triple-labeling and confocal microscopy, we tested the presence of TRPC1 by using a series of specific neurochemical markers. TRPC1 was abundant in SMI 32-positive pyramidal neurons, and in some glutamic acid decarboxylase 67 (GAD67) interneurons, but was lacking in glial fibrillary acidic protein (GFAP)-positive glial cells. In neurons it colocalized with postsynaptic marker MAP2 in cell bodies and apical dendritic trunks and it was virtually absent in synaptophysin-immunoreactive terminals. By using a panel of antibodies to classify interneurons, we identified the GABAergic interneurons that contained TRPC1. TRPC1 was lacking in basket and chandelier parvalbumin (PVALB) cells, and a very low percentage of calretinin (CALR) or calbindin (CALB) interneurons expressed TRPC1. Moreover, 63% of somatostatin (SST) expressing-cells and 37% of reelin-positive cells expressed TRPC1. All the SST/TRPC1 double-labeled cells, many of which were presumptive Martinotti cells (MC), were positive for reelin. The presence of TRPC1 in the somata and apical dendritic trunks of neocortical pyramidal cells suggests a role for this channel in sensory processing and synaptic plasticity. Conversely in SST/reelin interneurons, TRPC1 could modulate GABAergic transmission, which is responsible for shaping the coordinated activity of the pyramidal cells in the cortical network. In future studies, it would be relevant to investigate whether TRPC1 could be involved in the expression or processing of reelin in SST inhibitory interneurons.

Partial TrkB receptor activation suppresses cortical epileptogenesis through actions on parvalbumin interneurons

Post-traumatic epilepsy is one of the most common and difficult to treat forms of acquired epilepsy worldwide. Currently, there is no effective way to prevent post-traumatic epileptogenesis. It is known that abnormalities of interneurons, particularly parvalbumin-containing interneurons, play a critical role in epileptogenesis following traumatic brain injury. Thus, enhancing the function of existing parvalbumin interneurons might provide a logical therapeutic approach to prevention of post-traumatic epilepsy. The known positive effects of brain-derived neurotrophic factor on interneuronal growth and function through activation of its receptor tropomyosin receptor kinase B, and its decrease after traumatic brain injury, led us to hypothesize that enhancing trophic support might improve parvalbumin interneuronal function and decrease epileptogenesis. To test this hypothesis, we used the partial neocortical isolation (‘undercut’, UC) model of posttraumatic epileptogenesis in mature rats that were treated for 2 weeks, beginning on the day of injury, with LM22A-4, a newly designed partial agonist at the tropomyosin receptor kinase B. Effects of treatment were assessed with Western blots to measure pAKT/AKT; immunocytochemistry and whole cell patch clamp recordings to examine functional and structural properties of GABAergic interneurons; field potential recordings of epileptiform discharges in vitro; and video-EEG recordings of PTZ-induced seizures in vivo. Results showed that LM22A-4 treatment 1) increased pyramidal cell perisomatic immunoreactivity for VGAT, GAD65 and parvalbumin; 2) increased the density of close appositions of VGAT/gephyrin immunoreactive puncta (putative inhibitory synapses) on pyramidal cell somata; 3) increased the frequency of mIPSCs in pyramidal cells; and 4) decreased the incidence of spontaneous and evoked epileptiform discharges in vitro. 5) Treatment of rats with PTX BD4–3, another partial TrkB receptor agonist, reduced the incidence of bicuculline-induced ictal episodes in vitro and PTZ induced electrographic and behavioral ictal episodes in vivo. 6) Inactivation of TrkB receptors in undercut TrkBF616A mice with 1NMPP1 abolished both LM22A-4-induced effects on mIPSCs and on increased perisomatic VGAT-IR. Results indicate that chronic activation of the tropomyosin receptor kinase B by a partial agonist after cortical injury can enhance structural and functional measures of GABAergic inhibition and suppress posttraumatic epileptogenesis. Although the full agonist effects of brain-derived neurotrophic factor and tropomyosin receptor kinase B activation in epilepsy models have been controversial, the present results indicate that such trophic activation by a partial agonist may potentially serve as an effective therapeutic option for prophylactic treatment of posttraumatic epileptogenesis, and treatment of other neurological and psychiatric disorders whose pathogenesis involves impaired parvalbumin interneuronal function.

45. Rescue of the Functional Alterations of Motor Cortical Circuits in Arginase 1 Deficiency with AAV-Based Gene Therapy

The urea cycle is the main mechanism for terrestrial mammals to detoxify excess nitrogen. Disorders of the proximal urea cycle characteristically have periodic episodes of hyperammonemia leading to often severe and permanent neurological deterioration & disability. Ammonia has been implicated by compromising potassium buffering of astrocytic membranes and causing clinical neurological abnormalities by impairing cortical inhibition. Complete arginase 1 (Arg1) deficiency, a distal urea cycle disorder, is the least severe of these abnormalities, demonstrating neurological impairment including spasticity, loss of ambulation and seizures; while characterized by the presence of hyperargininemia, hyperammonemia is not a frequent clinical finding. While mortality is unfortunately common due to acute episodes of hyperammonemia in proximal urea cycle disorders, patients with hyperargininemia often are long-lived, however, suffering from progressive intellectual disability and spastic diplegia, and the mechanisms underlying the neurological dysfunction are not understood. To gain better insight on how the loss of arginase expression causes dysfunction in the developing brain, and if gene therapy could prevent these abnormalities, we studied how the excitability and functional and anatomical connectivity of motor cortical neurons are altered in the disorder using the murine knockout model. In addition, we examined if AAV expressing Arg1, administered IV on postnatal day 2, could rescue these findings. Results: Single- and double-copy loss of Arg1 caused dose-dependent decreases in intrinsic excitability, dendritic arborization complexity, and synapses in motor cortex layer V neurons. These findings show that 1) the intrinsic excitability of neurons of homozygous Arg1 knockout mice is abnormal and that, unexpectedly, heterozygous neurons (single copy loss) exhibit an intermediate phenotype compared to wild type and homozygous knockouts (double copy loss) (Fig. AFig. A); corresponding loss of Arg1 decreased the frequency of miniature excitatory postsynaptic currents and the amplitude of miniature inhibitory postsynaptic currents; 2) neuronal branching and spine phenotypes differ between genotypes with, unexpectedly, an intermediate phenotype for heterozygotes (Fig. BFig. B); and 3) with electron microscopic analysis and comparison of layer V synapses from arginase 1 knockout, heterozygous, and wild type mice, there is a very low density of excitatory (i.e. asymmetrical) synapses (Fig. CFig. C) in the knockout and decreased number of inhibitory (perisomatic) synapses (i.e. symmetrical) on somata of pyramidal cells, both dramatic findings. Finally, changes in synaptic morphology and abnormal ultrastructural features were found in knockout mice, also suggesting neuronal degeneration and inflammation. Neonatal intravenous administration on the second postnatal day with AAV expressing arginase 1 by a hepatocyte-specific promoter led to a near-resolution of these abnormalities when administered to homozygous Arg1 knockout animals. Summary: Our studies suggest that arginase 1 deficiency leads to severe and specific changes to intrinsic excitability and synaptic connectivity of motor cortical circuits. Importantly, we find that neonatal AAV-based Arg1 gene expression is effective in reversing both the physiological and anatomical hallmarks of the disorder.View Large Image | Download PowerPoint Slide

Mossy fiber sprouting and pyramidal cell dispersion in the hippocampal CA2 region in a mouse model of temporal lobe epilepsy.

Dentate granule cells and the hippocampal CA2 region are resistant to cell loss associated with mesial temporal lobe epilepsy (MTLE). It is known that granule cells undergo mossy fiber sprouting in the dentate gyrus which contributes to a recurrent, proepileptogenic circuitry in the hippocampus. Here it is shown that mossy fiber sprouting also targets CA2 pyramidal cell somata and that the CA2 region undergoes prominent structural reorganization under epileptic conditions. Using the intrahippocampal kainate mouse model for MTLE and the CA2-specific markers Purkinje cell protein 4 (PCP4) and regulator of G-Protein signaling 14 (RGS14), it was found that during epileptogenesis CA2 neurons survive and disperse in direction of CA3 and CA1 resulting in a significantly elongated CA2 region. Using transgenic mice that express enhanced green fluorescent protein (eGFP) in granule cells and mossy fibers, we show that the recently described mossy fiber projection to CA2 undergoes sprouting resulting in aberrant large, synaptoporin-expressing mossy fiber boutons which surround the CA2 pyramidal cell somata. This opens up the potential for altered synaptic transmission that might contribute to epileptic activity in CA2. Indeed, intrahippocampal recordings in freely moving mice revealed that epileptic activity occurs concomitantly in the dentate gyrus and in CA2. Altogether, the results call attention to CA2 as a region affected by MTLE-associated pathological restructuring.

Functional effects of distinct innervation styles of pyramidal cells by fast spiking cortical interneurons.

Inhibitory interneurons target precise membrane regions on pyramidal cells, but differences in their functional effects on somata, dendrites and spines remain unclear. We analyzed inhibitory synaptic events induced by cortical, fast-spiking (FS) basket cells which innervate dendritic shafts and spines as well as pyramidal cell somata. Serial electron micrograph (EMg) reconstructions showed that somatic synapses were larger than dendritic contacts. Simulations with precise anatomical and physiological data reveal functional differences between different innervation styles. FS cell soma-targeting synapses initiate a strong, global inhibition, those on shafts inhibit more restricted dendritic zones, while synapses on spines may mediate a strictly local veto. Thus, FS cell synapses of different sizes and sites provide functionally diverse forms of pyramidal cell inhibition.

Activity-dependent differences in function between proximal and distal Schaffer collaterals.

Axon conduction fidelity is important for signal transmission and has been studied in various axons, including the Schaffer collateral axons of the hippocampus. Previously, we reported that high-frequency stimulation (HFS) depresses Schaffer collateral excitability when assessed by whole-cell recordings from CA3 pyramidal cells but induces biphasic excitability changes (increase followed by decrease) in extracellular recordings of CA1 fiber volleys. Here, we examined responses from proximal (whole-cell or field-potential recordings from CA3 pyramidal cell somata) and distal (field-potential recordings from CA1 stratum radiatum) portions of the Schaffer collaterals during HFS and burst stimulation in hippocampal slices. Whole-cell and dual-field-potential recordings using 10-100-Hz HFS revealed frequency-dependent changes like those previously described, with higher frequencies producing more drastic changes. Dual-field-potential recordings revealed substantial differences in the response to HFS between proximal and distal regions of the Schaffer collaterals, with proximal axons depressing more strongly and only distal axons showing an initial excitability increase. Because CA3 pyramidal neurons normally fire in short bursts rather than long high-frequency trains, we repeated the dual recordings using 100-1,000-ms interval burst stimulation. Burst stimulation produced changes similar to those during HFS, with shorter intervals causing more drastic changes and substantial differences observed between proximal and distal axons. We suggest that functional differences between proximal and distal Schaffer collaterals may allow selective filtering of nonphysiological activity while maximizing successful conduction of physiological activity throughout an extensive axonal arbor.

VGluT3-expressing CCK-positive basket cells construct invaginating synapses enriched with endocannabinoid signaling proteins in particular cortical and cortex-like amygdaloid regions of mouse brains.

Invaginating synapses in the basal amygdala are a unique type of GABAergic synapses equipped with molecular-anatomical organization specialized for 2-arachidonoylglycerol (2-AG)-mediated endocannabinoid signaling. Cholecystokinin (CCK)-positive basket cell terminals protrude into pyramidal cell somata and form invaginating synapses, where apposing presynaptic and postsynaptic elements are highly loaded with cannabinoid receptor CB₁ or 2-AG synthetic enzyme diacylglycerol lipase-α (DGLα), respectively. The present study scrutinized their neurochemical and neuroanatomical phenotypes in adult mouse telencephalon. In the basal amygdala, vesicular glutamate transporter-3 (VGluT3) was transcribed in one-fourth of CB₁-expressing GABAergic interneurons. The majority of VGluT3-positive CB₁-expressing basket cell terminals apposed DGLα clusters, whereas the majority of VGluT3-negative ones did not. Importantly, VGluT3-positive basket cell terminals selectively constructed invaginating synapses. GABAA receptors accumulated on the postsynaptic membrane of invaginating synapses, whereas metabotropic glutamate receptor-5 (mGluR₅) was widely distributed on the somatodendritic surface of pyramidal cells. Moreover, CCK₂ receptor (CCK₂R) was highly transcribed in pyramidal cells. In cortical regions, pyramidal cells equipped with such VGluT3/CB₁/DGLα-accumulated invaginating synapses were found at variable frequencies depending on the subregions. Therefore, in addition to extreme proximity of CB₁- and DGLα-loaded presynaptic and postsynaptic elements, tripartite transmitter phenotype of GABA/glutamate/CCK is the common neurochemical feature of invaginating synapses, suggesting that glutamate, CCK, or both can promote 2-AG synthesis through activating Gαq/₁₁ protein-coupled mGluR₅ and CCK₂R. These molecular configurations led us to hypothesize that invaginating synapses might be evolved to provide some specific mechanisms of induction, regulation, and cooperativity for 2-AG-mediated retrograde signaling in particular cortical and cortex-like amygdaloid regions.

Role of small conductance Ca²⁺-activated K⁺ channels in controlling CA1 pyramidal cell excitability.

Small-conductance Ca(2+)-activated K(+) (SK or K(Ca)2) channels are widely expressed in the CNS. In several types of neurons, these channels were shown to become activated during repetitive firing, causing early spike frequency adaptation. In CA1 pyramidal cells, SK channels in dendritic spines were shown to regulate synaptic transmission. However, the presence of functional SK channels in the somata and their role in controlling the intrinsic firing of these neurons has been controversial. Using whole-cell voltage-clamp and current-clamp recordings in acute hippocampal slices and focal applications of irreversible and reversible SK channel blockers, we provide evidence that functional SK channels are expressed in the somata and proximal dendrites of adult rat CA1 pyramidal cells. Although these channels can generate a medium duration afterhyperpolarizing current, they play only an auxiliary role in controlling the intrinsic excitability of these neurons, secondary to the low voltage-activating, noninactivating K(V)7/M channels. As long as K(V)7/M channels are operative, activation of SK channels during repetitive firing does not notably affect the spike output of CA1 pyramidal cells. However, when K(V)7/M channel activity is compromised, SK channel activation significantly and uniquely reduces spike output of these neurons. Therefore, proximal SK channels provide a "second line of defense" against intrinsic hyperexcitability, which may play a role in multiple conditions in which K(V)7/M channels activity is compromised, such as hyposmolarity.

Cell type-specific expression of Eps8 in the mouse hippocampus

BackgroundEpidermal growth factor receptor substrate 8 (Eps8) is a multifunctional protein that regulates actin cytoskeleton dynamics and architecture through its barbed-end capping and bundling activities. In cultured hippocampal neurons, Eps8 is enriched at dendritic spine heads and is required for spine morphogenesis; however, the detailed expression pattern of Eps8 in the hippocampus has not yet been explored.ResultsHere, we demonstrate that endogenous Eps8 protein is restrictively expressed in neurons (NeuN-positive), but not in glial cells (glial fibrillary acidic protein-positive) in area CA1 of the mouse hippocampus. Surprisingly, Eps8 immunoreactivity is rarely found in pyramidal cell somata, but is expressed predominantly in the somata and dendrites of 67 kDa isoform of glutamic acid decarboxylase-expressing GABAergic interneurons in the stratum radiatum and at the border of stratum radiatum and lacunosum-moleculare of area CA1. On the basis of co-localizing markers, we found that Eps8 is not present in perisomatic inhibitory parvalbumin-expressing cells or calretinin-expressing interneurons. However, Eps8 is richly expressed in calbindin-expressing interneurons. Furthermore, Eps8 is also present in cholecystokinin-expressing interneurons, but not in somatostatin-expressing interneurons in area CA1 stratum pyramidale and stratum radiatum.ConclusionsThese results reveal a previously unknown cell type-specific expression pattern of endogenous Eps8 protein in the mouse hippocampus and speculate that the role of Eps8 in controlling and orchestrating neuronal morphogenesis and structural plasticity might be more prominent in interneurons than in pyramidal cells of the hippocampus.

Differential dendritic targeting of AMPA receptor subunit mRNAs in adult rat hippocampal principal neurons and interneurons

AbstractIn hippocampal neurons, AMPA receptors (AMPARs) mediate fast excitatory postsynaptic responses at glutamatergic synapses, and are involved in various forms of synaptic plasticity. Dendritic local protein synthesis of selected AMPAR subunit mRNAs is considered an additional mechanism to independently and rapidly control the strength of individual synapses.We have used fluorescent in situ hybridization and immunocytochemistry to analyze the localization of AMPAR subunit (GluA1–4) mRNAs and their relationship with the translation machinery in principal cells and interneurons of the adult rat hippocampus. The mRNAs encoding all four AMPAR subunits were detected in the somata and dendrites of CA3 and CA1 pyramidal cells and those of six classes of CA1 γ‐aminobutyric acid (GABA)ergic interneurons. GluA1–4 subunit mRNAs were highly localized to the apical dendrites of pyramidal cells, whereas in interneurons they were present in multiple dendrites. In contrast, in the dentate gyrus, GluA1–4 subunit mRNAs were virtually restricted to the somata and were absent from the dendrites of granule cells. These different regional and cell type‐specific labeling patterns also correlated with the localization of markers for components of the protein synthesis machinery. Our results support the local translation of GluA1–4 mRNAs in dendrites of hippocampal pyramidal cells and CA1 interneurons but not in granule cells of the dentate gyrus. Furthermore, the regional and cell type‐specific differences we observed suggest that each cell type uses distinct ways of regulating the local translation of AMPAR subunits. J. Comp. Neurol. 521:1954–2007, 2013. © 2012 Wiley Periodicals, Inc.

Differential dendritic targeting of AMPA receptor subunit mRNAs in adult rat hippocampal principal neurons and interneurons

In hippocampal neurons, AMPA receptors (AMPARs) mediate fast excitatory postsynaptic responses at glutamatergic synapses, and are involved in various forms of synaptic plasticity. Dendritic local protein synthesis of selected AMPAR subunit mRNAs is considered an additional mechanism to independently and rapidly control the strength of individual synapses. We have used fluorescent in situ hybridization and immunocytochemistry to analyze the localization of AMPAR subunit (GluA1-4) mRNAs and their relationship with the translation machinery in principal cells and interneurons of the adult rat hippocampus. The mRNAs encoding all four AMPAR subunits were detected in the somata and dendrites of CA3 and CA1 pyramidal cells and those of six classes of CA1 γ-aminobutyric acid (GABA)ergic interneurons. GluA1-4 subunit mRNAs were highly localized to the apical dendrites of pyramidal cells, whereas in interneurons they were present in multiple dendrites. In contrast, in the dentate gyrus, GluA1-4 subunit mRNAs were virtually restricted to the somata and were absent from the dendrites of granule cells. These different regional and cell type-specific labeling patterns also correlated with the localization of markers for components of the protein synthesis machinery. Our results support the local translation of GluA1-4 mRNAs in dendrites of hippocampal pyramidal cells and CA1 interneurons but not in granule cells of the dentate gyrus. Furthermore, the regional and cell type-specific differences we observed suggest that each cell type uses distinct ways of regulating the local translation of AMPAR subunits.

Multiple Layer 5 Pyramidal Cell Subtypes Relay Cortical Feedback from Secondary to Primary Motor Areas in Rats

Higher-order motor cortices, such as the secondary motor area (M2) in rodents, select future action patterns and transmit them to the primary motor cortex (M1). To better understand motor processing, we characterized "top-down" and "bottom-up" connectivities between M1 and M2 in the rat cortex. Somata of pyramidal cells (PCs) in M2 projecting to M1 were distributed in lower layer 2/3 (L2/3) and upper layer 5 (L5), whereas PCs projecting from M1 to M2 had somata distributed throughout L2/3 and L5. M2 afferents terminated preferentially in upper layer 1 of M1, which also receives indirect basal ganglia output through afferents from the ventral anterior and ventromedial thalamic nuclei. On the other hand, M1 afferents terminated preferentially in L2/3 of M2, a zone receiving indirect cerebellar output through thalamic afferents from the ventrolateral nucleus. While L5 corticopontine (CPn) cells with collaterals to the spinal cord did not participate in corticocortical projections, CPn cells with collaterals to the thalamus contributed preferentially to connections from M2 to M1. L5 callosal projection (commissural) cells participated in connectivity between M1 and M2 bidirectionally. We conclude that the connectivity between M1 and M2 is directionally specialized, involving specific PC subtypes that selectively target lamina receiving distinct thalamocortical inputs.

Cellular Targets of Nitric Oxide in the Hippocampus

In the hippocampus, as in many other CNS areas, nitric oxide (NO) participates in synaptic plasticity, manifested as changes in pre- and/or postsynaptic function. While it is known that these changes are brought about by cGMP following activation of guanylyl cyclase-coupled NO receptors attempts to locate cGMP by immunocytochemistry in hippocampal slices in response to NO have failed to detect the cGMP elevation where expected, i.e. in the pyramidal neurones. Instead, astrocytes, unidentified varicose fibres and GABA-ergic nerve terminals are reported to be the prominent NO targets, raising the possibility that NO acts indirectly via other cells. We have re-investigated the distribution of cGMP generated in response to endogenous and exogenous NO in hippocampal slices using immunohistochemistry and new conditions designed to optimise cGMP accumulation and, hence, its detectability. The conditions included use of tissue from the developing rat hippocampus, a potent inhibitor of phosphodiesterase-2, and an allosteric enhancer of the NO-receptive guanylyl cyclase. Under these conditions, cGMP was formed in response to endogenous NO and was found in a population of pyramidal cell somata in area CA3 and subiculum as well as in structures described previously. The additional presence of exogenous NO resulted in hippocampal cGMP reaching the highest level recorded for brain tissue (1700 pmol/mg protein) and in cGMP immunolabelling throughout the pyramidal cell layer. Populations of axons and interneurones were also stained. According with these results, immunohistochemistry for the common NO receptor β1-subunit indicated widespread expression. A similar staining pattern for the α1-subunit with an antibody used previously in the hippocampus and elsewhere, however, proved to be artefactual. The results indicate that the targets of NO in the hippocampus are more varied and extensive than previous evidence had suggested and, in particular, that the pyramidal neurones participating in NO-dependent synaptic plasticity are direct NO targets.

Temporal Dynamics of Parvalbumin-Expressing Axo-axonic and Basket Cells in the Rat Medial Prefrontal CortexIn Vivo

Axo-axonic interneurons, innervating exclusively axon initial segments, and parvalbumin-expressing basket interneurons, targeting somata, dendrites, and spines of pyramidal cells, have been proposed to control neuronal activity in prefrontal circuits. We recorded the spike-timing of identified neurons in the prelimbic cortex of anesthetized rats, and show that axo-axonic cells increase their firing during tail pinch-induced brain state-activation. In addition, axo-axonic cells differ from other GABAergic parvalbumin-expressing cells in their spike timing during DOWN- to UP-state transitions of slow oscillations and in their coupling to gamma and spindle oscillations. The distinct firing dynamics and synaptic targets of axo-axonic and other parvalbumin-expressing cells provide differential contributions to the temporal organization of prefrontal networks.

Quantitative Regional and Ultrastructural Localization of the Cav2.3 Subunit of R-type Calcium Channel in Mouse Brain

R-type calcium channels (RTCCs) are well known for their role in synaptic plasticity, but little is known about their subcellular distribution across various neuronal compartments. Using subtype-specific antibodies, we characterized the regional and subcellular localization of Ca(v)2.3 in mice and rats at both light and electron microscopic levels. Ca(v)2.3 immunogold particles were found to be predominantly presynaptic in the interpeduncular nucleus, but postsynaptic in other brain regions. Serial section analysis of electron microscopic images from the hippocampal CA1 revealed a higher density of immunogold particles in the dendritic shaft plasma membrane compared with the pyramidal cell somata. However, the labeling densities were not significantly different among the apical, oblique, or basal dendrites. Immunogold particles were also observed over the plasma membrane of dendritic spines, including both synaptic and extrasynaptic sites. Individual spine heads contained <20 immunogold particles, with an average density of ∼260 immunoparticles per μm(3) spine head volume, in accordance with the density of RTCCs estimated using calcium imaging (Sabatini and Svoboda, 2000). The Ca(v)2.3 density was variable among similar-sized spine heads and did not correlate with the density in the parent dendrite, implying that spines are individual calcium compartments operating autonomously from their parent dendrites.

Sticking out of the crowd: the molecular identity and development of cholecystokinin-containing basket cells

Certain essential cognitive processes require the precise temporal interplay between glutamatergic (excitatory) pyramidal cells and γ-aminobutyric acid (GABA)-releasing inhibitory interneurons in the hippocampus. Basket cells, the main class of interneurons, target pyramidal cell somata and proximal dendrites and thus are poised to modify network oscillations. Though only present in limited numbers, the impaired development of basket cells can result in changes in the hippocampal circuitry leading to neurological disorders, such as schizophrenia. The diversity of the spatial origins, neurochemical make-up, cytoarchitecture and network contributions amongst basket cells is a provocative example of interneuron heterogeneity in the hippocampus. This review discusses recent data concerned with the developmental trajectories of one subclass, the cholecystokinin-containing basket cell, and emphasizes the significance of the short-range intercellular guidance cues that have recently emerged to impact the formation and function of their inhibitory synapses.

Neural Activity and Neurotransmission Regulate the Maturation of the Innervation Field of Cortical GABAergic Interneurons in an Age-Dependent Manner

Neural activity guides the patterning of neuron synaptic territory in the developing nervous system. Evidence supporting this hypothesis comes from numerous studies on projection neurons in neuromuscular and visual systems. It is unknown whether the innervation field of GABAergic interneurons, which forms local dense innervations, follows similar rules. Cortical basket cells innervate hundreds of pyramidal cell somata and proximal dendrites. Thanks to this connectivity pattern, they can tightly control neural excitability and synchronization. Here we show that reducing excitation, and thus neurotransmitter release, in mouse cortical single basket cells in slice cultures decreases the number of innervated cells without changing the pattern of perisomatic innervation, both at the peak and after the proliferation phase of perisomatic synapse formation. Conversely, suppressing neurotransmitter release in single basket cells can have completely opposite effects depending on the developmental stage. Our results reveal a remarkably specific and age-dependent role of neural activity and neurotransmission levels in the establishment of the synaptic territory of cortical GABAergic cells.