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R Shigemoto

Publications and source records attributed to R Shigemoto.

At least 55 records · Page 3Linked to original sources

Metabotropic glutamate receptors are associated with non-synaptic appendages of unipolar brush cells in rat cerebellar cortex and cochlear nuclear complex.

Unipolar brush cells (UBCs) are a class of small neurons that are densely concentrated in the granular layers of the vestibulocerebellar cortex and dorsal cochlear nucleus. The UBCs form giant synapses with individual mossy fibre rosettes on the dendrioles which make up their brush formations and are provided with numerous, unusual non-synaptic appendages. In accord with the glutamatergic nature of mossy fibres, our previous post-embedding immunocytochemical studies indicated that various ionotropic glutamate receptor subunits are localized at the post-synaptic densities of the giant synapses, whereas the non-synaptic appendages are immunonegative. On the contrary, the metabotropic glutamate receptors mGluR1alpha and mGluR2/3 are situated at the non-synaptic appendages and are lacking at the post-synaptic densities. Other authors, however, have shown that antibodies to these metabotropic receptors stain both appendages and post-synaptic densities. In the present study, we have re-evaluated the distribution of metabotropic glutamate receptors in the UBCs of the cerebellum and the cochlear nuclear complex by light and electron microscopic pre-embedding immunocytochemistry with subtype-specific antibodies. We confirm that UBCs dendritic brushes are densely immunostained by antibody to mGluR1alpha particularly in the cerebellum and that antibody to mGluR2/3 labels at least a percentage of the UBC brushes in both the cerebellum and cochlear nuclei. At the ultrastructural level, it appears that mGluR1alpha and mGluR2/3 immunoreactivities are not associated with the post-synaptic densities of the giant mossy fibre-UBC synapses, but instead are concentrated on the non-synaptic appendages of the cerebellar UBCs. The non-synaptic appendages, therefore, may be an important avenue for regulating the excitability of UBCs and mediating glutamate effects on their still unknown intracellular signal transduction cascades. We also show that the pre-synaptic densities of UBC dendrodendritic junctions are mGluR2/3 positive. As previously demonstrated, antibodies to mGluR1alpha and mGluR2/3 label subsets of Golgi cells. Antibody to mGluR5 does not stain UBCs in the cerebellum and cochlear nucleus and reveals the somatodendritic compartment of Golgi cells situated in the core of the cerebellar granular layer, whilst cochlear nucleus Golgi cells are mGluR5 negative.

Animals↗

Collateral projections from striatonigral neurons to substance P receptor-expressing intrinsic neurons in the striatum of the rat.

It is well known that striatonigral neurons produce substance P (SP); however, no SP receptor (SPR) has so far been found in the substantia nigra. On the other hand, a previous study in the rat striatum indicated that SPR was expressed only in cholinergic or somatostatinergic intrinsic neurons (Kaneko et al. [1993] Brain Res. 631:297-303). Thus, it was assumed that SP produced by striatonigral neurons might be released through their intrastriatal axon collaterals to act upon intrinsic neurons in the striatum. To confirm this assumption, the distribution of axon collaterals of striatonigral neurons was examined in the striatum of the rat. The experiments were performed on brain slices by combining retrograde labeling with tetramethylrhodamine-dextran amine, electrophysiological recording, intracellular staining with biocytin, and immunocytochemistry for SPR. The distribution of axons of cholinergic striatal neurons (a group of SP-negative intrinsic striatal neurons) was also examined. It was observed that 16% of varicosities of intrastriatal axon collaterals of striatonigral neurons, as well as 6% of axonal varicosities of cholinergic neurons, were in close apposition to dendrites and cell bodies of SPR-immunoreactive striatal neurons. Since SPR-immunoreactive striatal neurons constituted only 2.7% of the total population of striatal neurons (Kaneko et al. [1993] Brain Res. 631:297-303), it appeared that axonal varicosities of striatonigral neurons were preferentially apposed to SPR-immunoreactive striatal neurons and that the varicosities in close apposition to SPR-immunoreactive neurons were derived more frequently from striatonigral neurons than from cholinergic interneurons. Confocal laser scanning microscopy indicated that axonal varicosities in close apposition to SPR-immunoreactive cells showed synaptophysin immunoreactivity, a marker of synaptic vesicles. In intrastriatal axons of striatonigral neurons, it was further revealed from electron microscopy that axonal varicosities in close apposition to SPR-immunoreactive dendrites, at least a part of them, made synapses of the symmetric type. Striatonigral neurons might release SP preferentially around cholinergic or somatostatinergic intrinsic neurons to regulate them through SP-SPR interactions.

Action Potentials↗

Differential presynaptic localization of metabotropic glutamate receptor subtypes in the rat hippocampus.

Neurotransmission in the hippocampus is modulated variously through presynaptic metabotropic glutamate receptors (mGluRs). To establish the precise localization of presynaptic mGluRs in the rat hippocampus, we used subtype-specific antibodies for eight mGluRs (mGluR1-mGluR8) for immunohistochemistry combined with lesioning of the three major hippocampal pathways: the perforant path, mossy fiber, and Schaffer collateral. Immunoreactivity for group II (mGluR2) and group III (mGluR4a, mGluR7a, mGluR7b, and mGluR8) mGluRs was predominantly localized to presynaptic elements, whereas that for group I mGluRs (mGluR1 and mGluR5) was localized to postsynaptic elements. The medial perforant path was strongly immunoreactive for mGluR2 and mGluR7a throughout the hippocampus, and the lateral perforant path was prominently immunoreactive for mGluR8 in the dentate gyrus and CA3 area. The mossy fiber was labeled for mGluR2, mGluR7a, and mGluR7b, whereas the Schaffer collateral was labeled only for mGluR7a. Electron microscopy further revealed the spatial segregation of group II and group III mGluRs within presynaptic elements. Immunolabeling for the group III receptors was predominantly observed in presynaptic active zones of asymmetrical and symmetrical synapses, whereas that for the group II receptor (mGluR2) was found in preterminal rather than terminal portions of axons. Target cell-specific segregation of receptors, first reported for mGluR7a (Shigemoto et al,., 1996), was also apparent for the other group III mGluRs, suggesting that transmitter release is differentially regulated by 2-amino-4-phosphonobutyrate-sensitive mGluRs in individual synapses on single axons according to the identity of postsynaptic neurons.

Animals↗

Relationship of NK3 receptor-immunoreactivity to subpopulations of neurons in rat spinal cord.

The distribution of immunoreactivity to the neurokinin3 receptor (NK3R) was examined in segments C7, T11-12, L1-2, and L4-6 of the rat spinal cord. NK3R immunoreactivity was visualized by using two antisera generated against sequences of amino acids contained in the C-terminal region of the NK3R. NK3R-immunoreactive cells were numerous in the substantia gelatinosa of all spinal segments examined as well as the dorsal commissural nucleus of spinal segments L1-2. Isolated, immunoreactive cells were scattered throughout other regions of the spinal cord. The relationship of NK3R-immunoreactivity with neurons was demonstrated by colocalization with microtubule associated protein 2-immunoreactivity in individual cells. Within neurons, NK3R-immunoreactivity was associated predominately with the plasma membrane of cell bodies and dendrites. Within the substantia gelatinosa, 86% of nitric oxide synthase (NOS)-immunoreactive neurons were also NK3R-immunoreactive. Although NOS-immunoreactive neurons were found throughout all other regions of the spinal cord in the segments examined, these were not NK3R-immunoreactive. When preganglionic sympathetic neurons in spinal segments T11-12 and L1-2 were visualized by intraperitoneal injection of Fluorogold, less than 1% of the Fluorogold-labeled neurons were also immunoreactive for NK3R. The large number of NK3R-immunoreactive neurons in the substantia gelatinosa suggests that some effects of tachykinins on somatosensation may be mediated by NK3R.

Animals↗

Localization of a metabotropic glutamate receptor, mGluR7, in axon terminals of presumed nociceptive, primary afferent fibers in the superficial layers of the spinal dorsal horn: an electron microscope study in the rat.

It was examined electron microscopically in the rat if a metabotropic glutamate receptor, mGluR7, might be localized in axon terminals of nociceptive, primary afferent fibers in laminae I and II of the spinal dorsal horn. Nociceptive nature of axon terminals showing mGluR7-like immunoreactivity (mGluR7-LI) was indicated by binding to the isolectin I-B4 from Griffonia simplicifolia (I-B4), or by substance P-like immunoreactivity (SP-LI). Axon terminals labeled with immunogold particles indicating mGluR7-LI were usually filled with round synaptic vesicles and were in asymmetric synaptic contact with dendritic or somatic profiles; occasionally they contained pleomorphic vesicles and were in symmetric synaptic contact with somatic profiles in lamina II. The double-labeling studies revealed that most of axon terminals with I-B4 labeling as well as a small population of axon terminals with SP-LI, showed mGluR7-LI. About one-third or much smaller population of axon terminals with mGluR7-LI in laminae I and II were labeled, respectively, with I-B4 or SP-LI; these were in asymmetric synaptic contact with dendritic profiles.

Animals↗

Distribution of trigeminohypothalamic and spinohypothalamic tract neurons displaying substance P receptor-like immunoreactivity in the rat.

Primary afferent neurons containing substance P (SP) are apparently implicated in the transmission of noxious information from the periphery to the central nervous system, and SP released from primary afferent neurons acts on second-order neurons with the SP receptor (SPR). In the rat, nociceptive information reached the hypothalamus not only through indirect pathways but also directly through trigeminohypothalamic and spinohypothalamic pathways. Thus, in the present study, the distribution pattern of trigeminohypothalamic and spinohypothalamic tract neurons showing SPR-like immunoreactivity (SPR-LI) was examined in the rat by a retrograde tract-tracing method combined with immunofluorescence histochemistry for SPR. A substantial number of trigeminal and spinal neurons with SPR-LI were retrogradely labeled with Fluoro-Gold (FG) injected into the hypothalamic regions. These neurons were distributed mainly in lamina I of the medullary and spinal dorsal horns, lateral spinal nucleus, regions around the central canal of the spinal cord, and the lateral aspect of the deep part of the spinal dorsal horn. A number of SPR-LI neurons in the spinal parasympathetic nucleus were labeled with FG injected into the area around the paraventricular hypothalamic nucleus. Some SPR-LI neurons in the lateral spinal nucleus and the lateral aspect of the deep part of the spinal dorsal horn were also labeled with FG injected into the septal region. On the basis of the distribution areas of SPR-LI trigeminal and spinal neurons projecting to the hypothalamic and septal regions, it is likely that these neurons are involved in the transmission of somatic and/or visceral noxious information.

Animals↗

Immunostaining for substance P receptor labels GABAergic cells with distinct termination patterns in the hippocampus.

A specific antiserum against substance P receptor (SPR) labels nonprincipal neurons in the cerebral cortex of the rat (T. Kaneko et al. [1994], Neuroscience 60:199-211; Y. Nakaya et al. [1994], J. Comp. Neurol. 347:249-274). In the present study, we aimed to identify the types of SPR-immunoreactive neurons in the hippocampus according to their content of neurochemical markers, which label interneuron populations with distinct termination patterns. Markers for perisomatic inhibitory cells, parvalbumin and cholecystokinin (CCK), colocalized with SPR in pyramidallike basket cells in the dentate gyrus and in large multipolar or bitufted cells within all hippocampal subfields respectively. A dense meshwork of SPR-immunoreactive spiny dendrites in the hilus and stratum lucidum of the CA3 region belonged largely to inhibitory cells terminating in the distal dendritic region of granule cells, as indicated by the somatostatin and neuropeptide Y (NPY) content. In addition, SPR and NPY were colocalized in numerous multipolar interneurons with dendrites branching close to the soma. Twenty-five percent of the SPR-immunoreactive cells overlapped with calretinin-positive neurons in all hippocampal subfields, showing that interneurons specialized to contact other gamma-aminobutyric acid-ergic cells may also contain SPR. On the basis of the known termination pattern of the colocalized markers, we conclude that SPR-positive interneurons are functionally heterogeneous and participate in different inhibitory processes: (1) perisomatic inhibition of principal cells (CCK-containing cells, and parvalbumin-positive cells in the dentate gyrus), (2) feedback dendritic inhibition in the entorhinal termination zone (somatostatin and NPY-containing cells), and (3) innervation of other interneurons (calretinin-containing cells).

Animals↗

Localisation of neurokinin 3 (NK3) receptor immunoreactivity in the rat gastrointestinal tract.

The localisation of the neurokinin 3 receptor (NK3r) in the rat gastrointestinal tract has been studied by using a polyclonal antiserum against the C-terminal portion (amino acids 388-452) of the rat NK3r. In the oesophagus, immunoreactivity for the NK3r was found on smooth muscle cells of the muscularis mucosae. NK3r immunoreactivity was not present on muscle cells of other regions. Nerve cell bodies immunoreactive for NK3r were seen in the myenteric and submucous plexuses of the small and large intestine, but not in the stomach or oesophagus. Immunoreactivity was largely confined to nerve cell surfaces. The reaction product was on the cell soma and initial parts of axons. Reactivity was not seen on nerve terminals. Immunoreactive nerve cells had Dogiel Type II morphology. Patterns of co-localisation of NK3r and immunoreactivity for other markers were examined in the ileum, to provide a basis from which to deduce the functional identity of NK3r-immunoreactive nerve cells. Most of the NK3r-immunoreactive nerve cells were also immunoreactive for the calcium-binding proteins, calretinin and calbindin, and all were immunoreactive for the NK1 receptor (NK1r). Nerve cells that were immunoreactive for nitric oxide synthase were not immunoreactive for either NK3r or NK1r. The projections of the calbindin and calretinin neurons were determined by nerve lesion studies. Their morphology, projections to the mucosa and other ganglia and immunoreactivity for the calcium-binding proteins suggest that the NK3r-immunoreactive neurons are intrinsic sensory neurons.

Animals↗

Differential plasma membrane distribution of metabotropic glutamate receptors mGluR1 alpha, mGluR2 and mGluR5, relative to neurotransmitter release sites.

Two group I metabotropic glutamate receptor subtypes, mGluR1 and mGluR5, have been reported to occur in highest concentration in an annulus surrounding the edge of the postsynaptic membrane specialisation. In order to determine whether such a distribution is uniform amongst postsynaptic mGluRs, their distribution was compared quantitatively by a pre-embedding silver-intensified immunogold technique at electron microscopic level in hippocampal pyramidal cells (mGluR5), cerebellar Purkinje cells (mGluR1 alpha) and Golgi cells (mGluR2). The results show that mGluR1 alpha, mGluR5 and mGluR2 each have a distinct distribution in relation to the glutamatergic synaptic junctions. On dendritic spines, mGluR1 alpha and mGluR5 showed the highest receptor density in a perisynaptic annulus (defined as within 60 nm of the edge of the synapse) followed by a decreasing extrasynaptic (60-900 nm) receptor level, but the gradient of decrease and the proportion of the perisynaptic pool (mGluR1 alpha, approximately 50%; vs mGluR5, approximately 25%) were different for the two receptors. The distributions of mGluR1 alpha and mGluR5 also differed significantly from simulated random distributions. In contrast, mGluR2 was not closely associated with glutamatergic synapses in the dendritic plasma membrane of cerebellar Golgi cells and its distribution relative to synapses is not different from simulated random distribution in the membrane. The somatic membrane, the axon and the synaptic boutons of the GABAergic Golgi cells also contained immunoreactive mGluR2 that is not associated with synaptic specialisations. In the hippocampal CA1 area the distribution of immunoparticles for mGluR5 on individual spines was established using serial sections. The results indicate that dendritic spines of pyramidal cells are heterogeneous with respect to the ratio of perisynaptic to extrasynaptic mGluR5 pools and about half of the immunopositive spines lack the perisynaptic pool. The quantitative comparison of receptor distributions demonstrates that mGluR1 alpha and mGluR5, but not mGluR2, are highly compartmentalised in different plasma membrane domains. The unique distribution of each mGluR subtype may reflect requirements for different transduction and effector mechanisms between cell types and different domains of the same cell, and suggests that the precise placement of receptors is a crucial factor contributing to neuronal communication.

Animals↗

The regional distribution and cellular localization of mRNA encoding rat prostacyclin synthase.

The cloned cDNA for rat prostacyclin synthase was found to contain a 1503-bp open reading frame which encoded a 501-amino acid protein sharing 84.0% identity with the human enzyme. RNA blot analysis revealed that the rat prostacyclin synthase mRNA, as a single species of 2.1 kb, is expressed abundantly in the aorta and uterus. High levels of expression were also observed in the stomach, lung, heart, testis, liver, and skeletal muscle. Low but significant expression was also seen in the brain and kidney. Furthermore, the regional distribution and cellular localization of prostacyclin synthase mRNA were examined by in situ hybridization analysis of rat tissue sections. The definitive signals for the mRNA were localized in smooth muscle cells of the arteries, bronchi and uterus, and in the cells of the fibrous tunic surrounding the seminiferous tubules, which are characterized as smooth muscle cells. Besides smooth muscle cells, signals were also detected in the fibroblasts of the heart myocardium, lung parenchyma cells and kidney inner medulla tubules and interstitial cells.

Amino Acid Sequence↗

Patchy distribution of substance P receptor immunoreactivity in the' developing rat striatum.

Developmental changes of the distribution pattern of substance P receptor (SPR) were investigated immunohistochemically in the rat striatum. The SPR immunoreactivity in the striatum first emerged at postnatal day 1 and transiently showed a patchy pattern of distribution until it displayed the adult pattern of homogeneous distribution by the third postnatal week. The SPR-immunoreactivity patches were most marked in the medial and dorsolateral parts of the striatum, as well as in the subcallosal streak. They matched tyrosine hydroxylase-enriched areas and, conversely, avoided calbindin-enriched zones. No neurons within the SPR-immunoreactive patches contained either choline acetyltransferase or somatostatin, which is known to be contained in intrinsic neurons in the striatum. The vast majority of SPR-immunoreactive patch neurons also contained DARPP-32, a phosphoprotein that is expressed in striatal projection neurons with D1 dopamine receptor. The results indicate that SPR-immunoreactive patches which appear transiently in the developing striatum are in register with the striatal patch compartment, and that SPR immunoreactivity within these patches may be expressed on projection neurons rather than intrinsic neurons. Such SPR immunoreactivity in projection neurons in striatal patches may fade out in adulthood.

Animals↗

Distribution of the mRNA for a pituitary adenylate cyclase-activating polypeptide receptor in the rat brain: an in situ hybridization study.

The distribution of the mRNA for a pituitary adenylate cyclase-activating polypeptide (PACAP) receptor (PACAP-R) was examined in the rat brain, and also in the hypophysis and pineal gland, by in situ hybridization with a specific 35S-labeled riboprobe which was generated from a rat PACAP-R cDNA clone. In the brain, expression of PACAP-R mRNA was most prominent in the periglomerular and granule cells of the olfactory bulb, granule cells of the dentate gyrus, supraoptic nucleus, and area postrema. The expression was also intense in the piriform, cingulate, and retrosplenial cortices, pyramidal cells in CA2, non-pyramidal cells in CA1-CA3, neuronal cells in the hilus of the dentate gyrus, lateral septal nucleus, intercalated amygdaloid nucleus, anterodorsal thalamic nucleus, most of the midline and intralaminar thalamic nuclei, many regions of the hypothalamus, dorsal motor nucleus of the vagus nerve, hypoglossal nucleus, and lateral reticular nucleus. No significant expression was detected in the mitral and tufted cells in the olfactory bulb, pyramidal cells in CA1 and CA3, posterior nuclear group of the thalamus, dorsal lateral geniculate nucleus, and Purkinje, Golgi, and granule cells in the cerebellar cortex. Moderate-to-weak expression was further observed in many other regions of the brain. In the cerebellar cortex, presumed Bergmann glia cells showed moderate expression. In the hypophysis, the expression was moderate in the anterior lobe, and weak to moderate in the posterior lobe; no significant expression was observed in the intermediate lobe. In the pineal gland, the expression was very weak, if any. Thus, the expression of PACAP-R was detected not only on neuronal cells but also on some particular glial cells. The present study has shown, for the first time, the exact site of PACAP-R expression in the brain and hypophysis. Although the functional significance of PACAP and PACAP-R in the brain still remains to be clarified, the present results are considered to provide some direction for future functional studies.

Animals↗

Impairment of hippocampal mossy fiber LTD in mice lacking mGluR2.

Subtype 2 of the metabotropic glutamate receptor (mGluR2) is expressed in the presynaptic elements of hippocampal mossy fiber-CA3 synapses. Knockout mice deficient in mGluR2 showed no histological changes and no alterations in basal synaptic transmission, paired-pulse facilitation, or tetanus-induced long-term potentiation (LTP) at the mossy fiber-CA3 synapses. Long-term depression (LTD) induced by low-frequency stimulation, however, was almost fully abolished. The mutant mice performed normally in water maze learning tasks. Thus, the presynaptic mGluR2 is essential for inducing LTD at the mossy fiber-CA3 synapses, but this hippocampal LTD does not seem to be required for spatial learning.

Animals↗

Target-cell-specific concentration of a metabotropic glutamate receptor in the presynaptic active zone.

The probability of synaptic neurotransmitter release from nerve terminals is regulated by presynaptic receptors responding to transmitters released from the same nerve terminal or from terminals of other neurons. The release of glutamate, the major excitatory neurotransmitter, is suppressed by presynaptic autoreceptors. Here we show that a metabotropic glutamate receptor (mGluR7) in the rat hippocampus is restricted to the presynaptic grid, the site of synaptic vesicle fusion. Pyramidal cell terminals presynaptic to mGluR1alpha-expressing interneurons have at least a ten-fold higher level of presynaptic mGluR7 than terminals making synapses with pyramidal cells and other types of interneuron. Distinct levels of mGluR7 are found at different synapses made by individual pyramidal axons or even single boutons. These results raise the possibility that presynaptic neurons could regulate the probability of transmitter release at individual synapses according to the postsynaptic target.

Animals↗

GABAergic synapses upon neurons expressing substance P receptors in the nucleus of the solitary tract: an immunocytochemical electron microscope study in the rat.

Morphological substrates for interactions between gamma-aminobutyric acid (GABA) and substance P upon neurons expressing substance P receptor (SPR) in the nucleus of the solitary tract (NST) were investigated by immunocytochemical electron microscopy. In the NST of the rat, many GABA-like immunoreactive axon terminals were in symmetric synaptic contacts with dendritic profiles; they were observed on nearly a half of the SPR-like immunoreactive dendritic profiles in the medial part of the caudal half of the NST.

Animals↗

Distribution of trigeminothalamic and spinothalamic-tract neurons showing substance P receptor-like immunoreactivity in the rat.

Trigeminothalamic and spinothalamic-tract neurons provided with substance P receptor (SPR) were examined in the rat by SPR immunofluorescence histochemistry combined with Fluoro-Gold (FG) fluorescent retrograde labeling. After FG injection in the thalamic regions, FG-labeled cells with SPR-like immunoreactivity were seen mainly in laminae I and III of the medullary and spinal dorsal horns and lateral spinal nucleus. In these regions, about one-fourth to one-third of FG-labeled cells showed SPR-like immunoreactivity.

Afferent Pathways↗

Presynaptic localization of a metabotropic glutamate receptor, mGluR4a, in the cerebellar cortex: a light and electron microscope study in the rat.

Localization of a metabotropic glutamate receptor, mGluR4a, was immunohistochemically examined in the rat cerebellum with an antibody, which was produced by using a synthetic peptide corresponding to a C-terminal sequence of rat mGluR4a. Marked mGluR4a-like immunoreactivity (mGluR4a-LI) was seen in neuropil of the molecular layer of the cerebellar cortex. Electron microscopically, mGluR4a-LI was observed in many axon terminals in the molecular layer. These axon terminals showing mGluR4a-LI were filled with round synaptic vesicles and were in asymmetric synaptic contacts most frequently with dendritic spines. The results indicate that mGluR4a are located presynaptically in the parallel fibers arising from the granule cells in the cerebellar cortex.

Animals↗

Presynaptic localization of a metabotropic glutamate receptor, mGluR8, in the rhinencephalic areas: a light and electron microscope study in the rat.

The present study indicated presynaptic localization of a metabotropic glutamate receptor, mGluR8, in projection neurons of the main olfactory bulb of rat. An antibody was produced by using a peptide corresponding to C-terminal 23 amino acids of mouse mGluR8. It was confirmed that the C-terminal 23 amino acids of rat mGluR8 were the same as those of mouse mGluR8 except for one, and that the antibody specifically recognized mGluR8 in the rat rhinencephalon. In layer Ia of the piriform cortex (a target area of projection fibers from the main olfactory bulb), mGluR8-like immunoreactivity (mGluR8-LI) was reduced after transection of the lateral olfactory tract, and mGluR8-LI was observed in axon terminals which were filled with round synaptic vesicles and made asymmetric synapses with dendritic spines.

Animals↗