Perivascular nerves with immunoreactivity to vasoactive intestinal polypeptide in cephalic arteries of the cat: distribution, possible origins and functional implications

The distribution of nerves containing vasoactive intestinal polypeptide(VIP)-immunoreactive material was examined in the cephalic arteries and cranial nerves of cats using an indirect immunofluorescence procedure on whole mounts. Perivascular VIP-immunoreactive nerves were widely distributed in arteries and arterioles supplying glands, muscles and mucous membranes of the face. Within the cerebral circulation, perivascular VIP-immunoreactive nerves were most abundant in the circle of Willis and the proximal portions of the major cerebral arteries and their proximal branches supplying the rostral brainstem and ventral areas of the cerebral cortex. Nerves containing VIP-immunoreactive material were absent from distal portions of arteries supplying the posterior brainstem, cerebellum and dorsal cerebral cortex. Cerebral perivascular VIP-immunoreactive nerves had extracerebral origins probably from VIP-immunoreactive perikarya within microganglia in the cavernous plexus and external rete. Extracerebral perivascular VIP-immunoreactive nerves probably arose from VIP-immunoreactive perikarya in microganglia associated with the tympanic plexus, chorda tympani, lingual nerve and Vidian nerve as well as from cells in the otic, sphenopalatine, submandibular and sublingual ganglia. Therefore, it seems likely that each major segment of the cephalic circulation is supplied by local VIP-immunoreactive neurons. If the VIP-immunoreactive nerves cause vasodilation, they are well placed to allow redistribution of arterial blood flow within the head. During heat stress, neurogenic vasodilation of the appropriate beds would permit efficient cooling of cerebral blood, particularly that supplying the rostral brainstem and surrounding areas of the cerebral cortex.     

Distribution of enteric neurons showing immunoreactivity for substance P in the guinea-pig ileum

Substance P-like immunoreactivity has been localized in whole mount preparations of the isolated layers of the guinea-pig ileum. Axons containing substance P formed dense networks around the nerve cells and ran in the primary, secondary and tertiary nerve bundles of the myenteric plexus. 3.6% of the nerve cell bodies of the myenteric plexus and 11.3% of the cell bodies in the submucous plexus showed immunoreactivity for substance P. Axons ran in fine nerve bundles parallel to the longitudinal muscle, between this muscle and the myenteric plexus. Axons containing substance P also ran in small nerve trunks parallel to the circular muscle throughout its thickness and in the deep muscular plexus at the base of this muscle coat. In the submucosa, these axons ramified amongst ganglion cells of the plexus and ran in the internodal strands. In addition they formed a perivascular network around submucous arteries and contributed to the paravascular nerves following these arteries. Axons containing substance P formed a delicate plexus in the mucosa. After extrinsic denervation the nerves containing substance P that were associated with submucous arteries, and some in the submucous plexus, disappeared. The nerves in the other areas were not detectably different from normal.Comparison with the distribution of somatostatin, enkephalin and vasoactive intestinal polypeptide indicated the neurons containing substance P constitute a separate population within the enteric nervous system.     

Synaptology and sources of vasoactive intestinal polypeptide and substance p containing axons of the cat celiac ganglion. An experimental electron microscopic immunohistochemical study

We have examined the possible origin of vasoactive intestinal polypeptide (VIP) and substance P-containing axons and the synapses formed by these axons in the celiac ganglion of the cat, by means of ultrastructural immunohistochemistry combined with various surgical lesions.Axons containing VIP as well as those immunopositive for substance P, formed axo-dendritic and axosomatic synapses with principal ganglion cells. After transaction of the superior mesenteric nerve numerous degenerated axonal profiles could be found in the celiac ganglion. Some of these contained VIP immunoreactivity. By contrast in alternate sections stained with substance P antibody only intact axons were labelled.Bilateral vagotomy resulted in the appearance of degenerating axonal profiles in the celiac ganglion, some of which could be stained with substance P antiserum but not with VIP antiserum. Following removal of dorsal root ganglia (Th₆-Th₁₂) from both sides, a large number of degenerated axons were found, many of which were immunopositive for substance P but not for VIP.We conclude that the peripheral input to the celiac ganglion contains VIP fibers which form synapses with principal ganglion cells. Substance P-containing fibers reach the celiac ganglion via the vagal nerve as well as from the dorsal root ganglia, and they form synapses with the principal ganglion cells. It is proposed that the VIP and the substance P-containing axons are involved in two different peripheral reflex loops.     

Origin of peptide-containing fibers in the inferior mesenteric ganglion of the guinea-pig: Immunohistochemical studies with antisera to substance P,enkephalin, vasoactive intestinal polypeptide,cholecystokinin and bombesin

After different denervation procedures the guinea-pig inferior mesenteric ganglion was analysed by immunohistochemistry using antisera to substance P, enkephalin, vasoactive intestinal polypeptide, cholecystokinin and bombesin. The results demonstrate that each of the nerve trunks connected to the ganglion carries specific peptidergic pathways. Thus, the lumbar splachnic nerves contain substance P-immunoreactive primary afferent neurons, which to a large extent traverse the ganglion, and enkephalin-immunoreactive preganglionic neurons; the colonie nerves carry vasoactive intestinal polypeptide-, cholecystokinin- and bombesin-immunoreactive fibers from the distal colon to the ganglion; the hypogastric nerves contain vasoactive intestinal polypeptide-positive fibers from the pelvic plexus; and the intermesenteric nerve contains vasoactive intestinal polypeptide, cholecystokinin, substance P and enkephalin from divergent sources. By studying accumulations of peptides in ligated lumbar splanchnic, intermesenteric, hypogastric and colonic nerves the existence of these major peptidergic pathways was confirmed and evidence was obtained for additional, not so prominent, peptidergic projections. The results are discussed in view of earlier morphological and physiological studies.     

Distribution of neuropeptides in the limbic system of the rat: the hippocampus

The distribution of several neuropeptides (vasoactive intestinal polypeptide, cholecystokinin octapeptide, substance P, neurotensin, methionine-enkephalin and somatostatin) in the hippocampal formation has been studied with immunocytochemical techniques. Numerous vasoactive intestinal polypeptide, cholecystokinin-octapeptide and somatostatin-positive cell bodies were found within the hippocampus and subiculum. Neurotensin-positive cell bodies were found within the subiculum, but no substance P or methionine-enkephalin-containing cell bodies were seen in either hippocampus proper or subiculum. Vasoactive intestinal polypeptide and cholecystokinin-octapeptide-positive cell bodies were predominantly located in the stratum moleculare and stratum radiatum of CA 1-2 regions and dentate gyrus, whilst somatostatin-containing cell bodies were found mainly in the stratum oriens. Nerve fibres containing each of the six peptides were found within the hippocampus. Large numbers of vasoactive intestinal polypeptide, cholecystokinin-octapeptide and somatostatin fibres innervated the pyramidal and granule cell layers, with smaller numbers in the stratum radiatum and fewer still in the stratum moleculare and stratum oriens. Other than a moderately dense neurotensin-positive fibre plexus in the dorsal subiculum, fewer neurotensin, substance P and methionine-enkephalin fibres were present. However, when present, these three peptides had a distribution restricted to a region close to the pyramidal layer in the CA 2/3 region and to the stratum moleculare of the CA 1 region. Peptide-containing fibres were identified entering or leaving the hippocampus in three ways, via (i) the fornix (all six peptides), (ii) the dorsal subiculum (neurotensin-positive fibres projecting to the cingulate cortex: somatostatin, vasoactive intestinal polypeptide, and cholecystokinin-octapeptide present in fibres running between the dorsal subiculum and occipito-parietal cortex) and (iii) the ventral subiculum (vasoactive intestinal polypeptide, cholecystokinin-octapeptide and somatostatin in fibres running between entorhinal cortex and hippocampus, and all six peptides in fibres crossing the amygdalo-hippocampal border). These findings indicate a major distinction between those peptides (vasoactive intestinal polypeptide, cholecystokinin-octapeptide, somatostatin, neurotensin) which are found in cell bodies intrinsic to the hippocampal formation and those peptides (substance P, methionine-enkephalin) which are found only in hippocampal afferents.(ABSTRACT TRUNCATED AT 400 WORDS)     

Distribution of neuropeptides in the limbic system of the rat: The amygdaloid complex

The distribution of six neuropeptides (vasoactive intestinal polypeptide, cholecystokinin octapeptide, substance P, neurotensin, methionine-enkephalin and somatostatin) has been mapped in the amygdala using immunocytochemical methods. Cell bodies containing each peptide showed a differential distribution throughout the various subnuclei. Large numbers of vasoactive intestinal polypeptide and cholecystokinin-octapeptide-containing cell bodies were located in the lateral and cortical nuclei respectively, neurotensin- and methionine enkephalin-containing cell bodies in the central nucleus, and substance P-containing cell bodies primarily in the medial nucleus. Somatostatin-containing cell bodies were found in all nuclei. Neuropeptide-containing fibres were also differentially distributed. Substance P and cholecystokinin fibres formed dense plexuses in the medial nucleus whilst the greatest concentration of vasoactive intestinal polypeptide, neurotensin and methionine enkephalin fibres were seen in the central nucleus. Close observation of serial sections showed that all the neuropeptides studied had extensive intra-amygdaloid pathways and connections with other brain areas.The central nucleus and stria terminalis have particular importance in the organisation of peptides within the amygdala. The central nucleus acts as a focus for a number of converging/diverging peptide pathways and incoming catecholaminergic afferents. The stria terminalis contains all six peptides and represents the major efferent peptidergic system. The amygdala is thought to control a number of endocrine reponses and to regulate complex behavioural functions. The abundance of neuropeptides within the amygdala and their complex pattern of pathways imply that they may act to regulate endocrine responses to external events (e.g. stress) or alter emotional tone, functions thought to be controlled by the amygdala.     

Coexistence of somatostatin- and avian pancreatic polypeptide (APP)-like immunoreactivity in some forebrain neurons

The indirect immunofluorescence technique was used to demonstrate the coexistence of somatostatin together with avian pancreatic polypeptide-like immunoreactivity within certain neurons of the rat forebrain. Numerous neurons containing these peptides were observed in the neocortex, hippocampus, olfactory tubercle, striatum, nucleus accumbens and lateral septum. In studies of serial sections stained alternately for these two peptides, and in restaining experiments, It could be determined that in many neurons in these areas these two peptides coexisted. In other brain areas such as the anterior periventricular hypothalamus, somatostatin cells were never found to contain avian pancreatic polypeptide-like immunoreactivity. Also, within the pancreas these two peptides were never found to coexist in the same cells. The findings represent a further example of the coexistence of more than one neuropeptide within a single neuron.     

Overall distribution of vasoactive intestinal polypeptide-containing nerves on the wall of cerebral arteries: an immunohistochemical study using whole-mounts

The overall distribution of vasoactive intestinal polypeptide (VIP)-like immunoreactivity on the wall of the cerebral arteries, including its 3-dimensional profile, was investigated by means of the indirect immunofluorescence method using flat-mounts. VIP-immunoreactive fibers run spirally on the wall of the cerebral arteries. On the wall of the large arteries, such as the vertebral artery, basilar artery, internal carotid artery, within and/or without the circle of Willis, posterior and anterior communicating arteries, proximal parts of anterior, mid and posterior cerebral arteries, these fibers are richly distributed and show a dense grid-like appearance. The highest density was identified on the wall of the anterior cerebral artery, internal carotid artery and anterior communicating artery, while the lowest density was on the posterior communicating artery. On the other hand, on the walls of the branches of these arteries or along distal parts of the anterior, mid and posterior cerebral arteries, the number of VIP-immunoreactive fibers decreased markedly.     

Effect of intrathecal capsaicin analogues on the immunofluorescence of peptides and serotonin in the dorsal horn in rats

The intrathecal administration of capsaicin, a homovanillylamide derivative, has been demonstrated to cause analgesia in response to thermal stimuli. This analgesia has been correlated with a profound depletion of spinal substance P, a putative primary afferent transmitter. We studied the effects of capsaicin, a series of capsaicin analogues, piperine and kainic acid on the immunohistochemical staining of substance P, cholecystokinin, somatostatin, methionine-enkephalin and serotonin. Capsaicin and an analogue 1-nonenoyl-vanillylamide significantly elevated the tail flick latency and when the spinal cords of the rats were analyzed immunohistochemically, a profound depletion of substance P and cholecystokinin was observed. The spinal somatostatin-immunoreactivity of these rats was slightly reduced. Piperine also depleted substance P and reduced somatostatin staining but did not alter the staining intensity or density of cholecystokinin, methionine-enkephalin or serotonin. Kainate-depleted methionine-enkephalin but did not alter any other neuropeptides studied or serotonin. These results may indicate a link between capsaicin-induced analgesia and the concomitant depletion of cholecystokinin and substance P.     

Distribution of neuropeptides in the limbic system of the rat: the bed nucleus of the stria terminalis, septum and preoptic area

The distribution of the neuropeptides vasoactive intestinal polypeptide, cholecystokinin octapeptide, substance P, neurotensin, methionine-enkephalin and somatostatin has been mapped immunocytochemically in the bed nucleus of the stria terminalis, one of the major sites of termination for efferent projections from the amygdala. Immunoreactive fibres and terminals were distributed more or less topographically and largely in accordance with the previously described localization of peptide-containing cell bodies in the amygdala and the amygdaloid projection fields in the bed nucleus as described by neuroanatomical techniques. Thus, vasoactive intestinal polypeptide, which was found in some of the lateral amygdaloid nuclei, had a substantial projection to the lateral bed nucleus. The lateral bed nucleus also contained cholecystokinin-octapeptide, substance P, neurotensin and methionine-enkephalin immunoreactivity which probably derived from the central amygdaloid nucleus, whilst cholecystokinin-octapeptide, and especially substance P-containing fibres, were found in the medial bed nucleus and probably arise from cells in the medial amygdala. Reciprocal amygdalopetal projections were suggested by the presence of substance P- and somatostatin-containing cell bodies in the mediodorsal bed nucleus and vasoactive intestinal polypeptide cells in the lateral bed nucleus, but somatostatin otherwise had a widespread distribution. Numerous local peptidergic connections were also noted both within the bed nucleus and between it and adjacent structures, especially the preoptic area, hypothalamus and the basal ganglia. These data provide further evidence that neuropeptides play a major role in the connectivity of the limbic system and show that the bed nucleus of the stria terminalis is an important relay station, particularly between amygdaloid efferents and other forebrain areas.     

The origins,pathways and terminations of neurons with VIP-like immunoreactivity in the guinea-pig small intestine

We have analyzed changes in the distributions of terminals with vasoactive intestinal polypeptide (VIP)-like immunoreactivity, and accumulations in severed processes, that occur after lesions of intrinsic and extrinsic nerve pathways of the guinea-pig small intestine. The observations indicate that enteric vasoactive intestinal polypeptide immunoreactive neurons have the following projections. Nerve cell bodies in the myenteric plexus provide varicose processes to the underlying circular muscle; the majority of these pathways, if they extend at all in the anal or oral directions, do so for distances of less than 1 mm. Nerve cell bodies of the myenteric plexus also project anally to provide terminals to other myenteric ganglia. The lengths of the majority of these projections are between 2 and 10 mm, with an average length of about 6 mm. Processes of myenteric neurons also run anally in the myenteric plexus and then penetrate the circular muscle to provide varicose processes in the submucous ganglia at distances of up to 15 mm, the average length being 9–12 mm. In addition, there is an intestinofugal projection of myenteric neurons whose processes end around nerve cell bodies of the coeliac ganglia. A similar projection from the colon supplies the inferior mesenteric ganglia. The nerve cell bodies in submucous ganglia give rise to a subepithelial network of fibres in the mucosa and also supply terminals to submucous arterioles.It is concluded that vasoactive intestinal polypeptide is contained in neurons of a number of intrinsic nerve pathways, influencing motility, blood flow and mucosal transport. The myenteric neurons that project to prevertebral sympathetic ganglia may be involved in intestino-intestinal reflexes.     

Neuronal configurations in lateral and basolateral amygdala

The lateral and basolateral nuclei of the rat amygdala have been studied with the rapid Golgi method. Both nuclei have similar cell types, which closely resemble cells in the cerebral cortex. Therefore, we suggest that what is known about cortical circuitry can be used as a guide for studying synaptic circuitry in the lateral and basolateral nuclei.

The most abundant neurons that are impregnated in both nuclei are pyramidal cells. They have conical cell bodies and easily recognizable apical and basilar dendrites. The ones in the center of each nucleus have long axes that roughly parallel the long axis of the nucleus. Towards the periphery, pyramidal cells have apical dendrites that either stick directly across the nucleus or follow along a nuclear border. The peripheral dendrites tend to enclose the nuclei. There is considerable overlap among the dendritic trees and the dendrites of one nucleus extend into the territory of the other. Pyramidal cells have extensive axonal systems. The principal axon of basolateral cells usually projects rostrally but long collaterals leave the nucleus in other directions. The axons of lateral nucleus pyramidal cells are also widely distributed. The major thrust of their axons is caudal and lateral.

Stellate cells are the most common variety of the non-pyramidal cells. They occur in both nuclei and have round cell bodies, 10–15 μm diameter, and spherical dendritic trees that are confined to a limited region of the nucleus. Their axons form dense terminal fields that remain within the vicinity of the parent cell's dendritic tree. Another type of non-pyramidal cell is the cone cell, whose non-spiny, varicose dendrites describe cones. These neurons are found mainly in the apex of the lateral nucleus. The most rare non-pyramidal cells are the extended neurons, which have long, straight dendrites that reach beyond the nucleus into surrounding neuropil. They are mostly in the rostral part of the basolateral nucleus but also occur in the lateral nucleus, near the ventricular border. The axons of cone cells and the extended neurons have been only partially impregnated.

We also have examined stellate cells in the guinea-pig lateral and basolateral nuclei. They have many of the same features as those in the rat brain, except that their dendritic trees and axonal systems are more complicated.

There are two large groups of afferents: one consists of longitudinally running axons and the other of transversely coursing fibers. The axons in both groups are thin, have a rather straight course through the nucleus and have only a few collaterals. We have attempted to relate these groups to the known afferent supply of the nuclei. In addition, axons from neurons in the intercalated cell masses and in the claustrum-dorsal endopiriform area enter at least the periphery of the lateral and basolateral nuclei.

Finally, we propose that the lateral and basolateral nuclei are functionally interrelated and that they link exteroceptive sensory data (‘percepts’) with the output of the basolateral nucleus, which is seen to coordinate forebrain areas that direct motivationally-guided behavior. Furthermore, this linked input-output pattern is communicated to the entorhinal cortex and, hence, to the hippocampus where it is encoded.     

Calcitonin gene-related peptide in cardiovascular tissues of the rat

The distribution of calcitonin gene-related peptide immunoreactivity in the cardiovascular system of the rat was investigated by radioimmunoassay and immunocytochemistry. The nature of the immunoreactivity was studied by gel permeation and high performance liquid chromatography. Immunocytochemistry demonstrated the existence of calcitonin gene-related peptide-containing nerve fibres throughout the cardiovascular system. These were present in all regions of the heart, particularly in association with the coronary arteries, within the papillary muscles and within the sinoatrial and atrioventricular nodes. Calcitonin gene-related peptide-containing fibres were found mainly in the adventitia of the arteries and veins. Calcitonin gene-related peptide concentrations were high in major arteries and veins but comparatively low in the heart, aortic arch and thoracic aorta. Chromatography showed that approximately 70% of the total immunoreactivity was identical to synthetic calcitonin gene-related peptide. Calcitonin gene-related peptide concentrations in the blood vessels of rats treated neonatally with capsaicin were not found to be significantly different from those in control animals although capsaicin caused significant reductions of calcitonin gene-related peptide levels in certain other tissues. The results of this study suggest that calcitonin gene-related peptide-containing fibres are likely to be of importance in the innervation of vascular tissues and raise the possibility that these fibres are different in character from calcitonin gene-related peptide-containing fibres found in other tissues.     

Distribution of enteric nerve cell bodies and axons showing immunoreactivity for vasoactive intestinal polypeptide in the guinea-pig intestine

Immunoreactivity for vasoactive intestinal polypeptide has been localized in neurons in the guinea-pig ileum, colon and stomach. In the ileum, 2.5% of the nerve cell bodies of the myenteric plexus and 45% of those of the submucous plexus showed vasoactive intestinal polypeptide-like immunoreactivity. Varicose axons containing vasoactive intestinal polypeptide ramified amongst the nerve cell bodies of both plexuses and in some cases formed rings of varicosities around non-reactive nerve cells. Axons were traced from the myenteric plexus to the circular muscle and deep muscular plexus. There were numerous positive axons running in fine strands within the circular muscle, parallel to the muscle bundles. Axons containing vasoactive intestinal polypeptide were associated with mucosal blood vessels, but few supplied the vascular network of the submucosa; some immunoreactive axons also contributed to the periglandular plexus of the mucosa. There were no changes in the distribution of axons in the ileum after extrinsic denervation.The results are discussed in relation to the possible functional roles of neurons that contain vasoactive intestinal polypeptide in the intestine: the distribution of such nerve cells in the myenteric plexus and of axons in the circular muscle and sphincters is consistent with this polypeptide being a transmitter of enteric inhibitory neurons; it is also possible that vasoactive intestinal polypeptide is the enteric vasodilator transmitter.     

P物质、血管活性肠肽和乙酰胆碱能神经在大鼠肠道内的分布及其关系

应用乙酰胆碱酯酶(AChE)组织化学和PAP免疫组织化学方法,比较观察P物质(SP)、血管活性肠肽(VIP)和AChE三种阳性神经元在大鼠十二指肠、空肠、回肠、结肠和直肠内的分布特征及其相互关系。结果显示:SP、VIP、AChE阳性神经神经元和纤维均分布于肠壁各层,从十二指肠、空肠到回肠逐渐增多,但从结肠到直肠则逐渐减少;AChE阳性神经元或纤维在肠壁各层最丰富,其中VIP以粘膜层和粘膜下神经丛较丰富,SP以肠肌丛较丰富;三者的分布密度为AChE>VIP>SP。AChE、SP和VIP阳性神经元胞体及神经纤维在不同肠段的分布密度有明显差异(P<0.05),提示可能与不同肠段肠动力调节功能有关。     

Substance P-containing sensory nerves in the rat iris. Normal distribution, ontogeny and innervation of intraocular iris grafts

Trigeminal, substance P-containing nerves have been studied in the stretch-prepared rat iris with immunohistochemical techniques. The normal iris exhibited a slightly irregular plexus of individual fibres in the dilator, intermingled with thin, meandering axon bundles. The sphincter contained more circumferentially oriented fibres. Occasional free nerve endings were present in all parts of the iris; no obvious association with blood vessels was detected. All substance P-positive nerves in the iris disappeared after lesioning the trigeminal nerve. Irides of neonates showed scattered, smooth fibres in a sparse plexus, without visible axon bundles. Over the first two postnatal weeks, the density of innervation developed rapidly, reaching a transiently supranormal level and fluorescence intensity, compared to adulthood. From 3 weeks on, the pattern and density of substance P-containing fibres approached the normal adult appearance. In irides grafted to the anterior eye chamber, the intrinsic substance P nerves degenerated, disappearing completely after 5 days. Reinnervation from the host irides transpired over the next few weeks, approximating normal density after 3 weeks, and organotypic density and distribution from 4 weeks on. No obvious hyperinnervation was encountered after longer postoperative times (3 months). In the host iris, many substance P fibres disappear or exhibit low fluorescence intensity during the first postoperative week, recovering fully during the next 2 weeks. Over longer postoperative periods irregular, moderate hyperinnervation developed with increased numbers of axons in bundles. In conclusion, we show normal distribution and plasticity during ontogeny and maturity of substance P-containing iris nerves in the rat, with a sensitive immunohistochemical technique in iris whole mounts.     

Bombesin-like immunoreactivity in sympathetic ganglia

Nerve fibres showing immunoreactivity for bombesin have been observed in prevertebral sympathetic ganglia of the rat and guinea-pig. The distribution of bombesin immunoreactive fibres was compared with the distribution of the structurally-related peptide substance P. Dense networks of varicose bombesin immunoreactive fibres were observed, and in the rat coeliac ganglion the density exceeded that of the substance P immunoreactive fibres. Extracts of rat and guinea-pig coeliac ganglion contained material cross-reacting in radioimmunoassays for bombesin tetradecapeptide. Similar concentrations of bombesin and substance P were found in the guinea-pig ganglion, whereas in the rat the concentration of bombesin was about 20 times as high as that of substance P. Two molecular forms of immunoreactive bombesin were identified, neither of which corresponded exactly to synthetic bombesin. The two components were also distinguishable from synthetic substance P. A potential role of a bombesin-like peptide in neural pathways between the gut and prevertebral ganglia is suggested.     

Cholecystokinin-like immunoreactive neurons in rat cerebral cortex

The distribution and form of cholecystokinin immunoreactive neurons in neocortical areas within the posterior pole of the rat cerebral hemisphere was examined using the immunoperoxidase technique. Although cholecystokinin-positive neurons are present throughout the cortex, they are most frequent in the supragranular layers. These neurons are of three kinds: layer I neurons, bipolar cells, and other non-pyramidal cells with either multipolar or bitufted dendritic trees. In electron-microscopic preparations, the horseradish peroxidase reaction product is found to form a granular deposit which occurs throughout the cytoplasm and nucleoplasm and shows no predilection for any particular type of organelle. Electron-microscopy also shows cholecystokinin-positive neurons to have both symmetric and asymmetric synapses on their perikarya, which is additional evidence in favor of the interpretation that they are non-pyramidal cells. In the light-microscopic preparations three types of CCK-positive axons are encountered. These are vertically-oriented axons considered to arise from bipolar cells, a plexus in the superficial portion of layer II/III which is believed to arise from the multipolar and bitufted cells, and a deep plexus of unknown origin in layers VI and V.Since the axons of bipolar cells form asymmetric synapses they are thought to be excitatory neurons. In contrast, the bitufted and multipolar neurons are probably inhibitory, for previous studies have shown neurons with similar features to have axons which form symmetric synapses and to contain glutamic acid decarboxylase. Thus, although iontophoretically-applied cholecystokinin excites cortical neurons, it appears to be present in some neurons which are excitatory and others which are inhibitory.