5-HT3 Receptors Mediate Serotonergic Fast Synaptic Excitation of Neocortical Vasoactive Intestinal Peptide/Cholecystokinin Interneurons

Neocortical neurons expressing the serotonin 5-HT3receptor (5-HT3R) were characterized in rat acute slices by using patch-clamp recordings combined with single-cell RT-PCR and histochemical labeling. The 5-HT3A receptor subunit was expressed selectively in a subset of GABAergic interneurons coexpressing cholecystokinin (CCK) and vasoactive intestinal peptide (VIP). The 5-HT3B subunit was never detected, indicating that 5-HT3Rs expressed by neocortical interneurons did not contain this subunit. In 5-HT3A-expressing VIP/CCK interneurons, serotonin induced fast membrane potential depolarizations by activating an inward current that was blocked by the selective 5-HT3R antagonist tropisetron. Furthermore, we observed close appositions between serotonergic fibers and the dendrites and somata of 5-HT3R-expressing neurons, suggestive of possible synaptic contacts. Indeed, in interneurons exhibiting rapid excitation by serotonin, local electrical stimulations evoked fast EPSCs of large amplitude that were blocked by tropisetron. Finally, 5-HT3R-expressing neurons were also excited by a nicotinic agonist, indicating that serotonergic and cholinergic fast synaptic transmission could converge onto VIP/CCK interneurons. Our results establish a clear correlation between the presence of the 5-HT3A receptor subunit in neocortical VIP/CCK GABAergic interneurons, its functional expression, and its synaptic activation by serotonergic afferent fibers from the brainstem raphe nuclei.

[1]  Alan Peters,et al.  Cellular components of the cerebral cortex , 1984 .

[2]  Kurochkina Ai,et al.  Classification of cortical neurons according to the character of their background impulse activity , 1975 .

[3]  R. S. Jones.  Responses of cortical neurones to stimulation of the nucleus raphé medianus: A pharmacological analysis of the role of indoleamines , 1982, Neuropharmacology.

[4]  P. Goldman-Rakic,et al.  Serotonergic axons in monkey prefrontal cerebral cortex synapse predominantly on interneurons as demonstrated by serial section electron microscopy , 1996, The Journal of comparative neurology.

[5]  J. Hornung,et al.  The Selective innervation by serotoninergic axons of calbindin‐containing interneurons in the neocortex and hippocampus of the marmoset , 1992, The Journal of comparative neurology.

[6]  Y. Kubota,et al.  GABAergic cell subtypes and their synaptic connections in rat frontal cortex. , 1997, Cerebral cortex.

[7]  E. Welker,et al.  Neurons immunoreactive for vasoactive intestinal polypeptide in the rat primary somatosensory cortex: Morphology and spatial relationship to barrel‐related columns , 2000, The Journal of comparative neurology.

[8]  P. Goldman-Rakic,et al.  Segregation of serotonin 5‐HT2A and 5‐HT3 receptors in inhibitory circuits of the primate cerebral cortex , 2000, The Journal of comparative neurology.

[9]  D. Wong,et al.  Localization of 5-HT3 receptors in the rat brain using [3H]LY278584 , 1991, Brain Research.

[10]  A. Brown,et al.  Ion permeation and conduction in a human recombinant 5‐HT3 receptor subunit (h5‐HT3A) , 1998, The Journal of physiology.

[11]  Pankaj Sah,et al.  Ca2+-activated K+ currents in neurones: types, physiological roles and modulation , 1996, Trends in Neurosciences.

[12]  Gavin Kilpatrick,et al.  Identification and distribution of 5-HT3 receptors in rat brain using radioligand binding , 1987, Nature.

[13]  B. Sakmann,et al.  Patch-clamp recordings from the soma and dendrites of neurons in brain slices using infrared video microscopy , 1993, Pflügers Archiv.

[14]  B. K. Hartman,et al.  Light and electron microscopic immunocytochemical analysis of the neurovascular relationships of choline acetyltransferase and vasoactive intestinal polypeptide nerve terminals in the rat cerebral cortex , 1994, The Journal of comparative neurology.

[15]  L. Acsády,et al.  Co-localization of vasoactive intestinal polypeptide, γ-aminobutyric acid and choline acetyltransferase in neocortical interneurons of the adult rat , 1997, Brain Research.

[16]  F. Bloom,et al.  The 5-HT3 Receptor Is Present in Different Subpopulations of GABAergic Neurons in the Rat Telencephalon , 1997, The Journal of Neuroscience.

[17]  G. Zupanc.  Peptidergic transmission: from morphological correlates to functional implications. , 1996, Micron.

[18]  P. Somogyi,et al.  Salient features of synaptic organisation in the cerebral cortex 1 Published on the World Wide Web on 3 March 1998. 1 , 1998, Brain Research Reviews.

[19]  J. DeFelipe,et al.  Synaptic relationships of serotonin-immunoreactive terminal baskets on GABA neurons in the cat auditory cortex. , 1991, Cerebral cortex.

[20]  Gavin Kilpatrick,et al.  The distribution of specific binding of the 5-HT3 receptor ligand [3H]GR65630 in rat brain using quantitative autoradiography , 1988, Neuroscience Letters.

[21]  H. Westenberg,et al.  Cholecystokinin in anxiety , 1996, European Neuropsychopharmacology.

[22]  K. Kawa,et al.  Distribution and functional properties of 5-HT3 receptors in the rat hippocampal dentate gyrus: a patch-clamp study. , 1994, Journal of neurophysiology.

[23]  J. Rossier,et al.  Kainate receptor subunits expressed in single cultured hippocampal neurons: Molecular and functional variants by RNA editing , 1995, Neuron.

[24]  J. A. van Hooft,et al.  Promiscuous coassembly of serotonin 5-HT3 and nicotinic alpha4 receptor subunits into Ca(2+)-permeable ion channels. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[25]  F. Bloom,et al.  Distribution of neurons expressing immunoreactivity for the 5HT3 receptor subtype in the rat brain and spinal cord , 1998, The Journal of comparative neurology.

[26]  D. McCormick,et al.  Comparative electrophysiology of pyramidal and sparsely spiny stellate neurons of the neocortex. , 1985, Journal of neurophysiology.

[27]  C R Houser,et al.  Morphological diversity of immunocytochemically identified GABA neurons in the monkey sensory-motor cortex , 1983, Journal of neurocytology.

[28]  F. Zhou,et al.  Activation of serotonin receptors modulates synaptic transmission in rat cerebral cortex. , 1999, Journal of neurophysiology.

[29]  J. Rossier,et al.  Classification of fusiform neocortical interneurons based on unsupervised clustering. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[30]  J. Rossier,et al.  Subunit composition at the single-cell level explains functional properties of a glutamate-gated channel , 1994, Neuron.

[31]  M. Mayer,et al.  Effects of 5-hydroxytryptamine on central neurones antagonized by bicuculline and picrotoxin , 1981, Neuropharmacology.

[32]  J. Parnavelas,et al.  Extensive co-existence of neuropeptides in the rat visual cortex , 1987, Brain Research.

[33]  L. Descarries,et al.  Ultrastructural relationships of serotonin axon terminals in the cerebral cortex of the adult rat , 1989, The Journal of comparative neurology.

[34]  R. North,et al.  5-hydroxytryptamine is a fast excitatory transmitter at 5-HT3 receptors in rat amygdala , 1992, Neuron.

[35]  Y. Kawaguchi,et al.  Groupings of nonpyramidal and pyramidal cells with specific physiological and morphological characteristics in rat frontal cortex. , 1993, Journal of neurophysiology.

[36]  R. Miledi,et al.  Inhibition of nicotinic acetylcholine receptors by bicuculline , 2001, Neuropharmacology.

[37]  R. North,et al.  5-HT3 receptors are membrane ion channels , 1989, Nature.

[38]  John A. Peters,et al.  The 5-HT3B subunit is a major determinant of serotonin-receptor function , 1999, Nature.

[39]  B. Hille,et al.  5‐HT3 receptor channels in dissociated rat superior cervical ganglion neurons. , 1992, The Journal of physiology.

[40]  E. Kirkness,et al.  Evidence for Expression of Heteromeric Serotonin 5‐HT3 Receptors in Rodents , 2000, Journal of neurochemistry.

[41]  M. Morales,et al.  Cellular and subcellular immunolocalization of the type 3 serotonin receptor in the rat central nervous system. , 1996, Brain research. Molecular brain research.

[42]  L. Descarries,et al.  Serotonin nerve terminals in adult rat neocortex , 1975, Brain Research.

[43]  G A Orban,et al.  Heterogeneity of GABAergic cells in cat visual cortex , 1988, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[44]  B. Costall,et al.  Characterisation and autoradiographic localisation of 5-HT3 receptor recognition sites identified with [3H]-(S)-zacopride in the forebrain of the rat , 1990, Neuropharmacology.

[45]  M. D. Goldfinger,et al.  Theoretical studies of impulse propagation in serotonergic axons , 1992, Biological Cybernetics.

[46]  M. Molliver,et al.  The serotonin innervation of the cerebral cortex in the rat—an immunohistochemical analysis , 1980, Neuroscience.

[47]  M. Hamon,et al.  Quantitative autoradiographic mapping of 5‐HT3 receptors in the rat CNS using [125I]iodo‐zacopride and [3H]zacopride as radioligands , 1992, Synapse.

[48]  F. Bloom,et al.  The type 3 serotonin receptor is expressed in a subpopulation of GABAergic neurons in the rat neocortex and hippocampus , 1996, Brain Research.

[49]  B. K. Hartman,et al.  Ultrastructural and morphometric features of the acetylcholine innervation in adult rat parietal cortex: An electron microscopic study in serial sections , 1994, The Journal of comparative neurology.

[50]  Y. Kubota,et al.  Three distinct subpopulations of GABAergic neurons in rat frontal agranular cortex , 1994, Brain Research.

[51]  J. Kauer,et al.  Hippocampal interneurons are excited via serotonin-gated ion channels. , 1997, Journal of neurophysiology.

[52]  I. Törk,et al.  Serotoninergic innervation of the cat cerebral cortex , 1988, The Journal of comparative neurology.

[53]  J. Yakel,et al.  The Nicotinic α4 Receptor Subunit Contributes to the Lining of the Ion Channel Pore When Expressed with the 5-HT3 Receptor Subunit* , 1999, The Journal of Biological Chemistry.

[54]  N. Ropert,et al.  Serotonin facilitates GABAergic transmission in the CA1 region of rat hippocampus in vitro. , 1991, The Journal of physiology.

[55]  F. E. Bloom,et al.  The distribution and morphological characteristics of the intracortical VIP-positive cell: An immunohistochemical analysis , 1984, Brain Research.

[56]  J. Rossier,et al.  AMPA receptor subunits expressed by single purkinje cells , 1992, Neuron.

[57]  J. Rossier,et al.  Properties of bipolar VIPergic interneurons and their excitation by pyramidal neurons in the rat neocortex , 1998, The European journal of neuroscience.

[58]  B. Connors,et al.  Intrinsic firing patterns of diverse neocortical neurons , 1990, Trends in Neurosciences.

[59]  E. G. Jones,et al.  Two classes of cortical GABA neurons defined by differential calcium binding protein immunoreactivities , 2004, Experimental Brain Research.

[60]  J. Rossier,et al.  Selective Excitation of Subtypes of Neocortical Interneurons by Nicotinic Receptors , 1999, The Journal of Neuroscience.

[61]  M. C. Angulo,et al.  Molecular and Physiological Diversity of Cortical Nonpyramidal Cells , 1997, The Journal of Neuroscience.

[62]  Y. Kubota,et al.  Physiological and morphological identification of somatostatin- or vasoactive intestinal polypeptide-containing cells among GABAergic cell subtypes in rat frontal cortex , 1996, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[63]  L. C. Katz,et al.  Fast Synaptic Signaling by Nicotinic Acetylcholine and Serotonin 5-HT3 Receptors in Developing Visual Cortex , 1997, The Journal of Neuroscience.

[64]  R. Myers,et al.  Primary structure and functional expression of the 5HT3 receptor, a serotonin-gated ion channel. , 1991, Science.

[65]  R. Buijs,et al.  Light and electron microscopic immunocytochemical analysis of the serotonin innervation of the rat visual cortex , 1987, Journal of neurocytology.

[66]  Yasuo Kawaguchi,et al.  Two distinct subgroups of cholecystokinin-immunoreactive cortical interneurons , 1997, Brain Research.

[67]  G. Papadopoulos,et al.  Serotoninergic afferents preferentially innervate distinct subclasses of peptidergic interneurons in the rat visual cortex , 2001, Brain Research.

[68]  P. Chomczyński,et al.  Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. , 1987, Analytical biochemistry.