Number Of Immunoreactive Neurons Research Articles

Persistence and atrophy of septal/diagonal band neurons expressing the p75 neurotrophin receptor in pilocarpine-induced chronic epilepsy in the rat

Systemic administration of pilocarpine, which results in status epilepticus followed by recurrent seizures in rats, is a widely used experimental model of chronic limbic epilepsy. Marked structural alterations have been documented in pilocarpine-induced epilepsy, and these include cell loss in the hippocampus and other brain areas, and sprouting of mossy and cholinergic fibers in the hippocampus. Evidence is accumulating that neurotrophins and neurotrophin receptors are involved in the cascade of these events. Two and 4 months after pilocarpine-induced epilepsy, neurons containing the 75-kDa low affinity neurotrophin receptor (p75 NTR) were investigated with immunohistochemistry in the medial septal and diagonal band nuclei. No significant differences in the distribution and number of immunoreactive neurons were found in the epileptic rats compared to control saline-treated animals. However, in the epileptic animals, a significant decrease in the perikaryal size of p75 NTR-immunoreactive neurons of the septal/diagonal band region was found by 60 days, and such atrophic changes were more marked in the diagonal band nuclei by 120 days. These findings indicate that the p75 NTR-containing cell bodies, which include the neurons projecting to the hippocampal formation and are cholinergic in the normal brain, survive after months of spontaneous recurrent seizures, during which, therefore, a supply of p75 NTR to target regions is maintained in the chronic epileptic brain. However, the present data point out that these p75 NTR-containing neurons undergo a significant shrinkage in pilocarpine-induced chronic epilepsy, thus indicating that they are involved in the brain pathology of temporal lobe epilepsy.

Excitability changes in freeze-induced neocortical microgyria.

A freezing probe was placed on the skull of postnatal day (PN) 1 rats to induce formation of a cerebrocortical microsulcus. Experimental studies were performed on PN days 21-24. At that time point, Nissl-stained sections revealed the presence of a microsulcus similar to that described in human dysplastic cortex. Immunocytochemical staining for parvalbumin, calretinin and calbindin indicated a significant decrease in the number of immunoreactive neurons within the microsulcus and non-significant decreases in regions adjacent to the microsulcus. Staining for the glial markers GFAP and vimentin was increased near the microsulcus. Using in vitro brain slices, recordings were made in cortex adjacent to the microsulcus. Epileptiform activity was observed in response to electrical stimulation near the microsulcus. Analysis of the voltage dependence of evoked epileptiform discharges suggested the presence of an inhibitory component. As previously observed in non-lesioned animals, bath application of 4-aminopyridine induced bicuculline-sensitive spontaneous burst discharges in the presence of excitatory amino acid antagonists. These results suggest that cortical freeze lesions associated with abnormal neuronal migration produce a chronic hyperexcitable state. The findings are consistent with a mechanism involving an alteration, not loss, of inhibition in this model.

Ultrastructural changes in nitric oxide synthase immunoreactivity in the brain of rats subjected to perinatal asphyxia: neuroprotective effects of cold treatment

Striatal and cortical neurons containing nitric oxide synthase (NOS) were studied in adult rats subjected to different periods of perinatal asphyxia (PA) using immunohistochemistry at both light microscopy (LM) and electron microscopy (EM). Another group was subjected to PA+hypothermia to study its neuroprotective effect. Quantitative image analysis was performed on the striatum and neocortex in order to count the number of immunoreactive neurons and to compare the pattern of staining between the different groups. Six-month-old rats that suffered subsevere and severe PA demonstrated, at LM, cytomegaly of the striatal and neocortical neurons containing NOS. Control and hypothermic neurons were more weakly immunostained than PA neurons. Subsevere and severe asphyctic rats showed an important neuronal loss that was reduced by hypothermic treatment. The PA group disclosed, at EM, dense electronic bodies distributed in terminals surrounding synaptic vesicles and in dendrites. Non-NOS-containing neurons showed signs of degeneration, such as dark cytoplasm and shrunken nuclei. Surrounding the blood vessels, we observed a clear edema. The immunolabeling in hypothermic rats resembled that observed in controls. These data suggest that subsevere and severe PA induces chronic changes in the neuronal content of NOS in the striatum and neocortex. Degeneration observed in neurons surrounding cytomegalic NOS-containing cells may be due to the excess of NO in their environment. Moreover, the chronic alterations produced by PA seem to be prevented by hypothermia.

Gradation of kainic acid-induced rat limbic seizures and expression of hippocampal heat shock protein-70.

Systemic injection of kainic acid (KA) induces limbic seizures in rats, which resemble human temporal lobe epilepsy, the most common form of adult human epilepsy. In this study, we have investigated KA-elicited limbic seizures in the rats by correlating the severity of the seizure attacks with the expression of hippocampal heat shock protein-70 (HSP70) which has been suggested to be a marker for neuronal injury/death in this model of seizures. After a systemic injection of KA, six stages of limbic seizures have been classified, namely, staring (stage 1), wet dog shake (stage 2), hyperactivity (stage 3), rearing (stage 4), rearing and falling (stage 5), and jumping (stage 6). Stages 4, 5 and 6 were further divided into mild and severe sub-stages. HSP70 expression was not detected in animals with stages 1 and 2 seizures. At stage 3 a small amount of HSP70 immunoreactive neurons was detected in the CA3 field and the dentate hilus. From stage 4 to stage 5 the degree of HSP70 immunoreactivity increased in the CA1 field from a few positive cells in stage 4 mild to large numbers of immunoreactive neurons in stage 5 severe. HSP70 became detectable in pyramidal cells in the CA2 field from stage 5 severe and higher. In animals with stage 6 seizures, the majority of HSP70 expression became located in glial cells throughout the whole hippocampus. We concluded that HSP70 expression in the hippocampus positively correlates with the severity of KA-elicited limbic seizures.

Oxidative Stress in Sudden Infant Death Syndrome

We investigated the hippocampus and parahippocampal cortex of victims of sudden infant death syndrome and of age-matched infants dying acutely of known causes (non-sudden infant death syndrome controls). Tissue sections were investigated for the presence of neurons expressing signs of elevated levels of free radical using immunohistochemical markers for superoxide dismutase and glutathione peroxidase. Brain tissues displayed immunopositive neurons in every infant. In control infants, an age-related decline in the number of superoxide dismutase- and glutathione peroxidase-immunoreactive neurons was apparent in the hippocampus and parahippocampal cortex. Significantly increased numbers of immunoreactive neurons were found in victims of sudden infant death syndrome under 6 months of age compared to age-matched controls. This suggests that infants who later become victims of sudden infant death syndrome may experience antemortem periods of oxidative stress, elevated levels of free radicals, and compensatory up-regulation of the free radical scavenger enzymes superoxide dismutase and glutathione peroxidase.

Co-localization of high-affinity neurotrophin receptors in nucleus basalis of Meynert neurons and their differential reduction in Alzheimer's disease

It has been suggested that degeneration of neurons in Alzheimer's disease is the result of diminished trophic support. However, so far no evidence has been forwarded that neuronal degeneration in Alzheimer's disease is causally related to insufficient production of neurotrophins. The present study deals with (i) the expression and co-localization of tyrosine kinase receptors (trks) in the human nucleus basalis of Meynert and (ii) alterations of these receptors in Alzheimer's disease in the nucleus basalis of Meynert, an area severely affected in Alzheimer's disease. The expression of trkA, trkB and trkC in the nucleus basalis of Meynert of control and Alzheimer's disease brains was studied using three polyclonal antibodies specifically recognizing the extracellular domain of trkA, trkB and trkC. Brain material of eight controls and seven Alzheimer's disease patients was obtained at autopsy, embedded in paraffin and stained immunocytochemically. Using an image analysis system, we determined the proportion of trk neurons expressing the different trk receptors in controls and Alzheimer's disease patients. In control brains, trkA, trkB and trkC were differentially expressed in numerous nucleus basalis of Meynert neurons. The highest proportion of neurons was found to express trkB (75%), followed by trkC (58%) and trkA (54%). Furthermore, using consecutive sections, a clear co-localization of trk receptors was observed in the same neurons. The highest degree of co-localization was observed between trkA and trkB. In Alzheimer's disease patients, the number of immunoreactive neurons and the staining intensity of individual neurons was reduced dramatically. Reduction in the proportion of neurons expressing trkA was 69%, in trkB 47% and in trkC 49%, which indicated a differential reduction in the amount of trk receptors in Alzheimer's disease.These observations indicate that nucleus basalis of Meynert neurons can be supported by more than one neurotrophin and that the degeneration of these neurons in Alzheimer's disease is associated with a decreased expression of trk receptors, suggesting a decreased neurotrophin responsiveness of nucleus basalis of Meynert neurons in Alzheimer's disease.

Reduced retrograde labelling with fluorescent tracer accompanies neuronal atrophy of basal forebrain cholinergic neurons in aged rats

During ageing, basal forebrain cholinergic neurons are prone to degeneration for unknown reasons. In this study we morphometrically evaluated the retrograde labelling of basal forebrain neurons obtained after injection of FluoroGold into multiple sites in the cerebral neocortex in aged (24–33 months) as compared with young adult (four to six months) male Sprague–Dawley rats. In addition, we looked for differences in the distribution of degenerative changes in topographic subdivisions of the basal forebrain cholinergic complex of neurons identified by immunohistochemical detection of the cholinergic markers choline acetyltransferase or low-affinity neurotrophin receptor. After injection of FluoroGold into the cerebral neocortex, the number of retrogradely labelled neurons in the horizontal diagonal band/substantia innominata and basal nucleus was significantly lower in aged rats, by 41% and 48%, respectively. In aged rats injected with FluoroGold, as well as in non-injected aged rats, the numbers of neurons immunoreactive for choline acetyltransferase and low-affinity neurotrophin receptor were significantly lower, by 23–27% in the basal forebrain system as a whole, with no significant difference in the degree of decline amongst different subdivisions (i.e. medial septum, diagonal band, substantia innominata and basal nucleus). The ratios of the number of neurons labelled with Fluoro Gold as compared with the number of neurons immunoreactive for either cholinergic marker were significantly lower in aged rats, by 32–37%, indicating that the decline in the number of neurons retrogradely transporting tracer was greater than the decline in the number of immunoreactive neurons in aged animals. Immunoreactive as well as retrogradely labelled neurons showed a significant shrinkage of cell surface area of 6–13% in different subdivisions of the basal forebrain cholinergic system in aged rats. These findings confirm significant loss and atrophy of basal forebrain cholinergic neurons in aged rats, and demonstrate significantly reduced retrograde labelling of these neurons with fluorescent tracer applied to their target cortex. This reduced retrograde labelling suggests an impairment of either uptake or retrograde transport mechanisms in these neurons in aged rats. Such an impairment may contribute to the degenerative changes of basal forebrain cholinergic neurons observed in ageing and age-related degenerative conditions such as Alzheimer's disease.

C-Fos expression in trigeminal nucleus neurons after chemical irritation of the cornea: reduction by selective blockade of nociceptor chemosensitivity.

The distribution and number of trigeminal brainstem and higher order sensory neurons expressing the protein product of the proto-oncogene c-fos after noxious stimulation of the cornea was studied in the rat using immunocytochemistry. The possibility that attenuation of nociceptive messages from the cornea by diltiazem reduced Fos-like immunoreactivity of spinal trigeminal neurons was also examined. A group of animals were killed 2-3 h after corneal stimulation. One cornea was stimulated with: a drop of 10 mM acetic acid; with acid plus mechanical scratching of the corneal epithelium; or with a drop of saline of 56 degrees C. Half of the animals treated with acid had been pretreated ipsilaterally with topical diltiazem (10 mM). Control rats received either saline in one eye or no treatment. Another group of animals were killed 7-8 h after stimulation with acetic acid. Fos-like immunoreactive neurons were counted in serial brainstem sections using an anti-Fos primary antiserum and processed according to the avidin-biotin complex method. In rats killed 2-3 h after corneal stimulation with acid, heat, or acid plus mechanical injury, labelled neurons were found in laminae I and II of the intermediate zone between caudalis and interpolaris subnuclei of the ipsilateral spinal trigeminal nucleus and, in a reduced number, in the symmetrical zones of the contralateral side. In animals stimulated with noxious heat or combined mechanical and chemical injury, a few scattered cells were also labelled in the ipsilateral junction between the cervical spinal cord and the caudalmost part of the trigeminal subnucleus caudalis. In rats killed 7 h after stimulation with acid, stained neurons were observed in the same areas of the trigeminal nucleus as in rats killed at shorter times, but in lower numbers; in these animals, no immunoreactive cells were found in deeper laminae or in higher sensory relay nuclei. Pretreatment with diltiazem significantly reduced the number of cells of the spinal trigeminal nucleus labelled after corneal stimulation with acid. The results indicate that brief noxious stimulation of the cornea evoke expression of c-Fos in neurons of the spinal trigeminal complex. Diminution by diltiazem of the number of immunoreactive neurons activated by corneal irritation suggests that this drug, by reducing chemosensitivity of nociceptive terminals, decreases nociceptive inflow to central nervous structures involved in ocular pain perception.

Polysialylated neural cell adhesion molecule expression by neurons and astrolial molecule in the rat dentate gyrus declines dramatically with increasing age

The expression of polysialylated neurons in the dentate gyrus of the hippocampal formation of young (postnatal day 40), mature (postnatal day 80) and aged (postnatal day 540) male Wistar rats has been investigated by immunohistochemical techniques employing a monoclonal antibody specific for neural cell adhesion molecule-linked α2,8 polysialic acid. A strong immunoreactivity was found on the cell bodies, dendrites and axons of granule-like neuronal cells at the border between the hilar region and the granule cell layer of the young rat. In the mature animal the number of immunoreactive neurons declined dramatically and were virtually absent in the aged group. Using an alternative fixation procedure, glial fibrillary acidic protein-positive and polysialylated astroglia processes were found in close proximity to the dendrites of the polysialylated granule-like cells. The number of astroglial processes traversing the granule cell layer showed a similar age-dependent decline to that observed with the polysialylated neurons. Glial fibrillary acidic protein-positive and polysialylated stellate astroglia were present throughout the hippocampal formation, but did not show the marked age-dependent decline observed with the astroglial processes in the granule cell layer.The neuronal dendrites and astroglial processes exhibited a strict numerical ratio in the young and mature animal and, in double immunofluorescence studies with anti-polysialic acid and anti-glial fibrillary acidic protein, the astroglial processes exhibited apparent points of cell and/or dendritic contact. These findings suggest that loss of polysialylated astroglial processes precedes the decline in polysialylated dentate neurons.

Comparison of c‐jun, junB, and junD mRNA expression and protein in the rat dorsal root ganglia following sciatic nerve transection

The present study was designed to compare the expression of the Jun family of protooncogenes following nerve injury. Adult rats were anesthetized and the sciatic nerve transected. Dorsal root ganglia (DRG) at 1, 2, 3, and 7 days after nerve transection were collected, their total RNA extracted, and Northern blots performed using 32P-labeled oligonucleotide probes. The constitutive expression of c-jun mRNA was very low in DRG. Induction of c-jun mRNA was observed by day 1 after nerve transection, with a sixfold peak at 3 days and a twofold induction still present by day 7. The constitutive expression of junB mRNA was also low in the DRG, and sciatic nerve transection produced only a modest induction (1.7-fold by day 3) in the DRG ipsilateral to the nerve cut. junD mRNA was constitutively expressed at high levels in the DRG, and its level of expression did not in the DRG, and its level of expression did not change after sciatic nerve transection. Immunocytochemistry studies demonstrated a pattern of c-Jun, JunB, and JunD immunoreactivity (IR) associated with the cell nuclei of DRG neurons. c-Jun IR was found at very low levels in the undamaged contralateral DRG neurons, but sciatic nerve transection dramatically increased the number of c-Jun-immunoreactive neurons. Dot blot immunoblotting assay confirmed that the DRG ipsilateral to the sciatic nerve cut contained a higher level of c-Jun protein than the contralateral control DRG. Similar to c-Jun IR, JunB IR was minimal in the undamaged contralateral DRG. However, the DRG ipsilateral to the nerve transection did not show an increase in the number of immunoreactive neurons. JunD protein was expressed at high levels in the contralateral DRG, and this level of expression persisted after sciatic nerve transection in the ipsilateral DRG. DNA gel retardation assay experiments with an AP-1 consensus sequence showed a single DNA-protein complex. This complex was increased in ipsilateral as compared with contralateral DRG extracts. The amount of DNA-protein complex was reduced by c-Jun protein antiserum but was not altered when treated with a Fos antibody. We conclude that c-jun, junB and junD mRNAs and proteins are differentially regulated in the DRG after sciatic nerve transection.

Increase in alpha-CGRP and GAP-43 in aged motoneurons: a study of peptides, growth factors, and ChAT mRNA in the lumbar spinal cord of senescent rats with symptoms of hindlimb incapacities.

Sprague-Dawley rats develop progressive motor dysfunctions during the third year of life. We use this as a model to examine possible neuronal mechanism(s) that may cause motor impairments occuring during aging. In this study we have used indirect immunofluorescence histochemistry (IF) and in situ hybridization histochemistry (ISH) to study quantitatively and qualitatively the staining pattern and mRNA expression of calcitonin gene-related peptide (alpha-CGRP), growth-associated protein 43 (GAP-43), and acidic fibroblast growth factor (aFGF) in spinal lumbar motoneurons of young adult (2-3 months) and aged (30 months) Sprague-Dawley rats. In addition, mRNAs encoding choline acetyltransferase (ChAT), beta-CGRP, and cholecystokinin (CCK) were analyzed. All aged rats used in this study disclosed symptoms of hindlimb incapacity, ranging from mild weight-bearing insufficiency to paralysis of the hind limbs. The symptoms were confined to the musculature of the hindlimb and hip regions. Only a small number (approximately 15%) of the large motoneurons that innervate the hindlimb muscles were lost in those aged rats that had clinical symptoms of hindlimb motor incapacities. The remaining motoneurons expressed ChAT mRNA at levels similar to those of young adult rats. The vast majority of these motoneurons showed increased mRNA levels for alpha-CGRP and GAP-43. Aged motoneurons contained more CGRP like immunoreactivity (LI), but the number of immunoreactive neurons was smaller than in adult rats. GAP-43-LI could be detected in motoneurons in aged, but not in adult, rats. GAP-43-LI was always colocalized with CGRP-LI in aged motoneurons. Studies of individual aged rats revealed that the increase of GAP-43 mRNA-positive cell bodies occurred in cases with the most severe clinical symptoms, whereas the increase in alpha-CGRP was even evident in rats with mild symptoms. No alterations in content of aFGF-LI or aFGF mRNA could be detected in the aged rat, and the content of CCK and beta-CGRP mRNAs was also normal. The usefulness of this rat model for studies of neuromuscular aging and possible functional roles for GAP-43 and CGRP in plastic and regenerative processes during aging are discussed.

Inhibition of axoplasmic transport in the rat vagus nerve alters the numbers of neuropeptide and tyrosine hydroxylase messenger RNA-containing and immunoreactive visceral afferent neurons of the nodose ganglion.

Previous work showed that axotomy-induced deafferentation of the placode-derived visceral afferent neurons of the nodose ganglion altered their expression of some neuropeptides and tyrosine hydroxylase. The present studies were designed to selectively evaluate the loss of axonal transport on the numbers of vasoactive intestinal polypeptide, tyrosine hydroxylase, and calcitonin gene-related peptide mRNA-containing and immunoreactive neurons in the nodose ganglion of the adult rat. Vinblastine (0.15 mM) application to the cervical vagus nerve was used to block axonal transport between ganglionic perikarya and peripheral targets. In situ hybridization histochemistry with 35S-labeled oligonucleotide probes was used to both quantify the number of mRNA-containing neurons and to assess the density of mRNA expression per neuron, and immunocytochemistry was used to visualize the number of immunoreactive neurons. The efficacy of vinblastine to inhibit axonal transport was verified by evaluating the build-up of calcitonin gene-related peptide immunoreactive in the vagus nerve immediately rostral to the site of drug application. The absence of vinblastine-induced neuronal damage was verified by the relative absence of degenerating nerves in the vagus nerve caudal to the site of drug application. Vinblastine treatment of the vagus nerve increased the numbers of vasoactive intestinal peptide mRNA-containing neurons and vasoactive intestinal peptide-immunoreactive neurons in the nodose ganglion at three, seven and 14 days, and increased the numbers of calcitonin gene-related peptide mRNA-containing and calcitonin gene-related peptide-immunoreactive neurons in the nodose ganglion at one, three and seven days. The average labeling density of vasoactive intestinal peptide mRNA-containing neurons was also increased following vinblastine treatment. Vinblastine treatment of the cervical vagus nerve, however, led to the appearance of low-labeling density calcitonin gene-related peptide mRNA-neurons and resulted in reduction of the average labeling density for calcitonin gene-related peptide mRNA-containing neurons. In contrast, application of vinblastine to the cervical vagus nerve, decreased the number of tyrosine hydroxylase mRNA-containing and tyrosine hydroxylase-immunoreactive neurons in the nodose ganglion. In summary, inhibition of the axoplasmic transport between the periphery and the visceral sensory perikarya appeared to alter vasoactive intestinal peptide, calcitonin gene-related peptide, and tyrosine hydroxylase expression and content in visceral sensory neurons of the nodose ganglion. These data suggest the presence of an axonally transported influence on the regulation of neuropeptide and neurotransmitter enzyme synthesis in mature placode-derived visceral sensory neurons.

Somatostatin and leu-enkephalin in the rat auditory brainstem during fetal and postnatal development.

A transient expression of the neuropeptide somatostatin has been described in several brain areas during early ontogeny and several opioid peptides, such as leu-enkephalin, have also been found in the brain at this stage in development. It is therefore believed that somatostatin and leu-enkephalin may play a role in neural maturation. The aim of the present study was to describe the spatiotemporal pattern of somatostatin and leu-enkephalin immunoreactivity in the auditory brainstem nuclei of the developing rat and to correlate it with other developmental events. In order to achieve this goal, we applied peroxidase-antiperoxidase immunocytochemistry to rat brains between embryonic day (E) 17 and adulthood. Somatostatin immunoreactivity (SIR) was found in all nuclei of the auditory brainstem, yet it was temporally restricted in most nuclei. SIR appeared prenatally and reached maximum levels around postnatal day (P) 7, when great numbers of immunoreactive neurons were present in the ventral cochlear nucleus (VCN) and in the lateral lemniscus. At that time relatively low numbers of cells were labeled in the dorsal cochlear nucleus, the lateral superior olive (LSO), and the inferior colliculus (IC). During the same period, when somata in the VCN were somatostatin-immunoreactive (SIR), a dense network of labeled fibers was also present in the LSO, the medial superior olive (MSO), and the medial nucleus of the trapezoid body (MNTB). As these nuclei receive direct input from VCN neurons, and as the distribution and morphology of the somatostatinergic fibers in the superior olivary complex (SOC) was like that of axons from VCN neurons, these findings suggest a transient somatostatinergic connection within the auditory system. Aside from the LSO, MSO, and MNTB, labeled fibers were found to a smaller extent in all other auditory brainstem nuclei. After P7, the SIR decreased and only a few immunoreactive elements were found in the adult auditory brainstem nuclei, indicating that somatostatin is transiently expressed in the rat auditory brainstem. Leu-enkephalin immunoreactivity showed a lower number and weaker intensity of labeled structures as compared to SIR, with E18 being the earliest day at which labeled fibers appeared in the SOC. At birth, immunoreactive fibers were also present in the cochlear nuclear complex and in the IC. Leu-enkephalin immunoreactive somata were found only after P12 in the CN and after P16 in the IC. Leu-enkephalin immunoreactivity was not transient, but increased progressively with age until about P21, when the adult levels were reached.(ABSTRACT TRUNCATED AT 400 WORDS)

Localization of neuropeptides in the nervous system of the brittle star Ophiura ophiura

Immunocytochemical investigations using antisera against the SALMFamide neuropeptide S1 and RFamide were carried out on whole mounts of the radial nerve cord and circumoral ring of the brittle star Ophiura ophiura. Both antisera show an abundant immunoreactivity in the ectoneural tissue and give similar patterns of distribution. They show that in the radial nerve cord there are discrete populations of neurons organized in an identical pattern in each segment. At the junction with the ring, however, the two proximal segments of the nerve cord are differentiated from the distal ones by increased numbers of immunoreactive neurons. This is probably due to the more complex integrative function in that area. In the ring, immunolabelling is simple and consists of fibre tracts and two local ganglia. In each segment both antisera label either one or two giant neurons the shape of which is similar to that of a class of fibres that have been described from intra-cellular dyefills. The S1- and RFamide-like distributions, although very similar, show some neurons which are only labelled by one antiserum. Since preabsorption controls indicate no cross-reactivity between antisera, there is evidence of two distinct neuropeptides in the nervous system of the brittle star.

Reappraisal of the innervation of rat intestine by vasoactive intestinal polypeptide and neuropeptide Y-immunoreactive neurons

The occurrence and distribution of neurons and nerve fibres showing vasoactive intestinal polypeptide-like and neuropeptide Y-like immunoreactivity were re-examined in the enteric nervous system of the small and large intestine of the adult rat using dual-labelling indirect immunofluorescence histochemistry to detect the co-existence of these neuropeptides. In the myenteric plexus of both small and large intestine, a population of neuropeptide Y-immunoreactive neurons that did not contain vasoactive intestinal polypeptide was noted; it accounted for 29–53% of neuropeptide Y neurons. Such neurons were also found in the submucosa but there they constituted at most 2% of neuropeptide Y-immunoreactive neurons. In both myenteric and submucous plexuses, regional variations were observed in the number of immunoreactive neurons and in the proportion of dual-labelled neurons. In the myenteric plexus, for example, the density of neurons with immunoreactivity to these two neuropeptides was constant throughout the small intestine, whereas it progressively increased distally within the colon. In addition, a distinct but small subset of immunoreactive myenteric neurons was found to have a novel soma morphology, unclassifiable according to the criteria used for porcine or guinea-pig enteric neurons. Such neurons had one or more conspicuous processes, which were much longer than the short, lamellar somal processes of typical Dogiel Type I neurons; moreover, these protruded from an essentially smooth soma and terminated at distances of up to two cell diameters from their point of origin. Thus, our results suggest that the organization of the enteric nervous system of the rat differs from that of other species and indicate that investigations of the co-localizations of neuropeptides and biologically active mediators in the intestinal tract would be incomplete without reference to regional differences in the incidence and distribution of such neurochemicals.

Localization of leucokinin-like immunoreactive neurons and their metamorphic changes in the ventral ganglion of the fly, Sarcophaga bullata

The distribution of leucokinin I-like immunoreactive neurons in the ventral ganglion of the fly Sarcophaga bullata was examined by indirect immunofluorescence. In the larval ventral ganglion there are seven pairs of large, highly immunoreactive neurons distributed ventrolaterally as bilateral pairs in abdominal neuromeres 1–7. During metamorphosis, the seven pairs of larval immunoreactive neurons survive and three additional pairs of immunoreactive neurons appear within the condensed abdominal ganglion, bringing the total number of immunoreactive neurons to 10 pairs. It is suggested that the neuropeptide from the newly formed three pairs of leucokinin-like immunoreactive neurons may have some unique function in the life of the adult insect.

Vasoactive intestinal polypeptide (VIP)-containing neurons: distribution throughout the brain of the chick (Gallus domesticus) with focus upon the lateral septal organ.

The distribution of VIP-like perikarya and fibers was determined throughout the chick brain. The most rostral immunoreactive perikarya were found to be cerebrospinal fluid-contacting neurons in the pars medialis of the lateral septal organ. Additional data were presented supporting the idea that the lateral septal organ is another circumventricular organ within the brain of birds (Kuenzel and van Tienhoven 1982). A large group of immunoreactive perikarya was found in the lateral hypothalamic area and appeared continuous with immunoreactive neurons in the anterior medial and ventromedial hypothalamic nuclei (n). A few perikarya were located in the paraventricular hypothalamic n. A number of immunoreactive neurons were found within and about the infundibular and inferior hypothalamic n., none however was immunoreactive cerebrospinal fluid-contacting neurons. Immunoreactive perikarya were found predominantly in laminae 10-11 of the stratum griseum et fibrosum superficiale. A few scattered perikarya were found ventromedial to the n. tegmenti pedunculo-pontinus pars compacta and locus ceruleus. Some of the immunoreactivity was unusual, being very homogeneous within the cell body with little evidence of the material in the axon or dendrites. Perikarya were found in the central gray, n. intercollicularis, and area ventralis of Tsai. The most caudal structure showing immunoreactive neurons was the n. reticularis paragigantocellularis lateralis. Brain areas containing the most abundant immunoreactive fibers, listed from the rostral-most location, were found in the ventromedial region of the lobus parolfactorius and the lateral septal n. Continuing caudally, there were immunoreactive fibers within the periventricular hypothalamic n.; some of the fibers were found to travel for some distance parallel to the third ventricle. Dense immunoreactive fibers were found in the tractus cortico-habenularis et cortico-septalis, medial habenular n. and posterior and dorsal n. of the archistriatum. A number of areas had what appeared to be baskets of immunoreactive fibers (perhaps immunoreactive terminals) surrounding non-reactive perikarya. Brain areas containing terminals included the piriform cortex, area ventralis of Tsai, interpeduncular n., and specific regions of the stratum griseum et fibrosum superficiale. A very dense immunoreactivity occurred within the external zone of the median eminence, the dorsolateral parabrachial n., and n. tractus solitarii. Vasoactive intestinal polypeptide appears to be a useful peptide for defining the neuroanatomical constituents of the visceral forebrain in birds.

Distribution, morphological features, and synaptic connections of parvalbumin- and calbindin D28k-immunoreactive neurons in the human hippocampal formation.

Calcium binding proteins calbindin D28k (CaBP) and parvalbumin (PV) are known to form distinct subpopulations of gamma-aminobutyric acid (GABA)ergic neurons in the rodent hippocampal formation. Light and electron microscopic morphology and connections of these protein-containing neurons are only partly known in the primate hippocampus. In this study, CaBP and PV were localized in neurons of the human hippocampal formation including the subicular complex (prosubiculum, subiculum, and presubiculum) in order to explore to what extent these subpopulations of hippocampal neurons differ in phylogenetically distant species. CaBP immunoreactivity was present in virtually all granule cells of the dentate gyrus and population of in a proportion of pyramidal neurons in the CA1 and CA2 regions. A distinct population of CaBP-positive local circuit neurons was found in all layers of the dentate gyrus and Ammon's horn. Most frequently they were located in the molecular layer of the dentate gyrus and the pyramidal layer of Ammon's horn. In the subicular complex pyramidal neurons were not immunoreactive for CaBP. In the prosubiculum and subiculum immunoreactive nonpyramidal neurons were equally distributed in all layers, whereas in the presubiculum they occurred mainly in the superficial layers. Electron microscopy showed typical somatic and dendritic features of the granule, pyramidal, and local circuit neurons. CaBP-positive mossy fiber terminals in the hilus of the dentate gyrus and terminals of presumed pyramidal neurons of Ammon's horn formed asymmetric synapses with dendrites and spines. CaBP-positive terminals of nonprincipal neurons formed symmetric synapses with dendrites and dendritic spines, but never with somata or axon initial segments. PV was exclusively present in local circuit neurons in both the hippocampal formation and subicular complex. Most of the PV-positive cell bodies were located among or close to the principal cell layers. However, large numbers of immunoreactive neurons were also found in the molecular layer of the dentate gyrus and in strata oriens of Ammon's horn. PV-positive cells were equally distributed in all layers of the subicular complex. Electron microscopy showed the characteristic somatic and dendritic features of local circuit neurons. PV-positive axon terminals formed exclusively symmetric synapses with somata, axon initial segments and dendritic shafts, and in a few cases with dendritic spines. The CaBP- and PV-containing neurons formed similar subpopulations in rodents, monkeys, and humans, although the human hippocampus displayed the largest variability of these immunoreactive neurons in their morphology and location. Calcium binding protein-containing neurons frequently occurred in the molecular layer of the human dentate gyrus and in the stratum lacunosum-moleculare of Ammon's horn.(ABSTRACT TRUNCATED AT 400 WORDS)

Measurements of Somatostatin and Neuropeptide Y in the Visual Cortex of Monocularly Deprived Rats

The levels of somatostatin and neuropeptide Y were measured with radioimmunoassay bilaterally in visual cortical areas 17, 18, and 18a of rats which had received monocular enucleation at birth. Neuropeptide levels were consistently three- to fourfold higher for neuropeptide Y than for somatostatin. Monocular enucleation did not change somatostatin levels within areas 17 or 18 of either hemisphere but significantly increased somatostatin levels in contralateral area 18a when compared to contralateral areas 17 or 18 3 months after enucleation. The concentrations of neuropeptide Y are significantly greater in areas 17 and 18a than those in area 18, however, neonatal enucleation had no significant effect on neuropeotide Y levels within any visual cortical area of either hemisphere. Visual cortical areas 17, 18, and 18a show differences in the relative concentrations of neoropeptide Y compared to somatostatin. Furthermore, these two peptides respond distinctively to neonatal enucleation. Enuclestion had no effect on the concentration of either peptide in samples of frontal cortex. Immunohistochemical analysis showed that area 17 contains far fewer somatostatin neurons than areas 18 or 18a, in marked contrast to the uniform levels of somatostatin measured in all visual cortical areas by radioimmunoassay. Immunohistochemically identified neuropeptide Y-immunoreactive neurons are evenly distributed between areas 17, 18, and 18a and represent about half of the number of somatostatin-immunoreactive cells. While neuropeptide Y levels are significantly different between these visual cortical areas, the numbers of immunoreactive neurons are similar. Thus, relatively few neuropeptide Y cells are accompanied by neuropeptide Y concentrations that are four- to fivefold higher than those for somatostatin, the more abundant cell type. In view of the high degree of colocalization of neuropeptide Y and somatostatin in cortical nonpyramidal neurons, these observations imply differences in the biosynthetic and degradation/release mechanisms in these neurons.

Distribution of prosaposin‐like immunoreactivity in rat brain

Prosaposin is the precursor for saposins A, B, C, and D, which are small lysosomal proteins required for the hydrolysis of sphingolipids by specific lysosomal hydrolases. With a monospecific anti-saposin C antibody, which cross-reacts with prosaposin but not with saposin A, B, or D, the present immunoblot experiments showed that the rat brain expresses an unprocessed approximately 72 kDa protein (possibly prosaposin) and little saposin C. Regional analysis demonstrated that prosaposin is abundant in the brainstem, hypothalamus, cerebellum, striatum, and hippocampus, and less abundant in the cerebral cortex. Consistent with this finding, prosaposin-like immunoreactive neurons and fibers as revealed by immunohistochemistry were observed frequently in subcortical regions. The medial septum, diagonal bands, basal nucleus of Meynert, ventral striatum, medial habenular nucleus, and motor nuclei of cranial nerve had significant numbers of immunoreactive neurons. There were also nerve fibers with prosaposin-like immunoreactivity in several projection fields of the above nuclei. Other brain areas that contained prosaposin-like immunoreactive neurons and/or processes were: several brain nuclei (nucleus caudate putamen, globus pallidus, substantia nigra, red nucleus) constituting the so-called extrapyramidal system, reticular thalamic nucleus, entopeduncular nucleus, mammillary nuclei, auditory relay nuclei, cerebellum, sensory cranial nerve nuclei, and the reticular formation. The distribution pattern of prosaposin is apparently different from that of other neuroactive substances so far examined, and thus prosaposin may be involved in novel central events.