TH-like Immunoreactivity Research Articles

Immunohistochemical investigation of nerve distribution in mature parotid and submandibular glands of rats with a liquid diet.

Although feeding with a liquid diet does not affect the growth of rat submandibular glands, it inhibits the growth of rat parotid glands during growth periods. In these growth-inhibited parotid glands, the growth of parasympathetic nerves is also suppressed. Meanwhile, the mature parotid glands of animals maintained on a liquid diet become morphologically and functionally atrophic, however, there is no effect of a liquid diet on mature submandibular glands. The objective of the present study was to clarify whether the nerve distribution in the mature salivary glands of rats was affected by a liquid diet. Seven-week-old male Wistar rats were used in this study. Half of the rats were kept on a pellet diet, and half were kept on a liquid diet, for 3, 7, 14, or 21 days. All rats were euthanised by isoflurane at each endpoint. Then, the parotid and submandibular glands were removed, frozen in liquid nitrogen, cryosectioned, and stained with antibodies against protein gene product 9.5 (PGP 9.5; general nerve marker), tyrosine hydroxylase (TH; sympathetic nerve marker), or neuronal nitric oxide synthase (nNOS; parasympathetic nerve marker). In parotid and submandibular glands of the pellet diet group, PGP 9.5- and TH-like immunoreactivity were seen around acini and ducts, and nNOS-like immunoreactivity was lower than PGP 9.5- and TH-like immunoreactivity. In the parotid glands of the liquid diet group, similar immunoreactivities were seen throughout the experimental period. The distribution of antibody labelling in the submandibular glands of the liquid diet group was similar to that of the pellet diet group and remained unchanged during the experimental period. The present study demonstrated no regressive effects of a liquid diet on the distribution of sympathetic or parasympathetic nerves in mature parotid glands and submandibular glands. This differed from inhibitory effects on the growth of parotid glands seen during growth periods.

Tyrosine hydroxylase in mouse pancreatic islet cells, in situ and after syngeneic transplantation to kidney.

Tyrosine hydroxylase (TH) is co-expressed with islet hormones in the fetal mouse pancreas. In the adult animal, the enzyme has been considered as a marker of ageing beta-cells. By immunohistochemical staining, we analyzed the expression of TH-like immunoreactivity (TH-LI), insulin-LI (INS-LI) and somatostatin-LI (SOM-LI) in adult mouse islets, in situ and after isolation and transplantation to kidney. In pancreas in situ, most TH-LI cells expressed INS-LI while less than 5% expressed SOM-LI. The total number of TH-LI cells/mm2 was significantly increased directly after isolation and in 0-day, 12-week and 52-week old grafts, but not in 3-day grafts. The proportion of TH-LI cells expressing SOM-LI increased after transplantation, amounting to about one-third by 52 weeks. As expressed per unit islet area, the frequencies of both TH/INS and TH/SOM cells increased significantly in the transplants. The results demonstrate that TH occurs in both beta-cells and D-cells of adult islets. In both cell types the enzyme appears to be responsive to the microenvironmental changes inherent in transplantation. This cellular phenotype plasticity might contribute to the altered insulin secretory dynamics in islet grafts.

Fos expression in catecholaminergic medullary neurons induced by chemical stimulation of stomach projecting to the paraventricular nucleus of the hypothalamus in rats.

To determine whether medullary catecholaminergic neurons expressing Fos induced by chemical stimulation of the stomach project to the paraventricular nucleus of hypothalamus (PVH) in rats. Horseradish peroxidase (HRP) was introduced stereotaxically into the PVH of rats. Histochemical analysis of coronal sections through the medulla were analyzed using triple-label immunohistochemistry to identify cells that were retrogradely labeled with HRP, Fos (ABC method), and tyrosin hydroxylase (TH) (PAP method). Seven kinds of labeled neurons were found in the nucleus tractus solitarii (NTS), the ventrolateral medulla (VLM) and the reticular formation (RF) of the medulla: Fos-like immunoreactive (FosLI) neurons, TH-like immunoreactive (TH-LI) neurons and HRP retrogradely single-labeled neurons, FosLI/HRP, FosLI/TH-LI and HRP/TH-LI double-labeled neurons, and FosLI/HRP/TH-LI triple-labeled neurons. Ascending projections from the NTS, VLM and RF to the PVH might be involved in the transmitting process of visceral noxious stimulation.

Evidence of early nervous differentiation and early catecholaminergic sensory system during Sepia officinalis embryogenesis

Within Mollusca, cephalopods exhibit a particularly complex nervous system. The adult brain is formed from the fusion of several "typical" molluscan ganglia but it remains poorly understood how these ganglia emerge, migrate, and differentiate during embryogenesis. We studied the development of both central and peripheral nervous system by antibodies raised against alpha-tubulin and tyrosine hydroxylase (TH) in Sepia officinalis embryos to visualize neurites and catecholamine-containing neurons, respectively. In early embryos, when organs start delineating, some ganglia already exhibited a significant fiber network. TH-like immunoreactivity was detected in these fibers and in some primary sensory neurons in the embryo periphery. These data attest to the occurrence of an early embryonic sensory nervous system, likely effective, transient in part, and in relation to the perception of external cues. Concerning the peripheral nervous network, the stellate ganglia emerged as a plexus of numerous converging axons from TH-like immunoreactive sensory cells, first at the mantle edge, and then in the whole mantle surface. Later, TH-immunopositive motor fibers, originating from the stellate ganglia, penetrated the circular muscles of the mantle. These patterns reveal the setup of a mantle midline with likely attractive and repulsive properties. Our findings seem to challenge the widespread, still accepted, view of a late differentiation of cephalopod ganglia, and provides significant data for further investigations about axonal guidance during cephalopod development.

Projections to the alimentary canal from the dopaminergic neurons in the dorsal motor nucleus of the vagus of the rat

The motility of the alimentary canal is regulated not only by neurons that contain acetylcholine or adrenaline, but also by nonadrenergic noncholinergic neurons. There are many neurons containing dopamine in the dorsal motor nucleus of the vagus (DMV). We examined the projections of these dopaminergic neurons to the alimentary canal with double-labeling immunohistochemistry for tyrosine hydroxylase (TH) and the retrograde tracer cholera toxin subunit b following its injection into the subdiaphragmatic esophagus, the cardia, the pylorus, the duodenum, the jejunum, and the ascending colon. Almost all double-labeled neurons were found in the half of the DMV caudal to the area postrema. In the caudal half of the DMV, about 58% of the TH-immunoreactive neurons projected to the cardia, about 36% projected to the pylorus, and about 28% projected to the subdiaphragmatic esophagus. Only a few TH-immunoreactive neurons projected to the duodenum, the jejunum, or the ascending colon. As a whole, less than 10% of the neurons in the DMV that projected to the alimentary canal showed TH-like immunoreactivity. These results suggest that some of the dopaminergic neurons in the DMV might regulate the activities of the stomach and the subdiaphragmatic esophagus.

“Blood‐contacting neurons” in the brain of the Japanese eel Anguilla japonica

To discriminate "blood-contacting neurons" within the brain of the eel, Evans blue (EB) was injected intraperitoneally. After five days, six brain areas were externally stained blue with the dye; the saccus dorsalis (SD), the epiphysis (E), the area postrema (AP), the posterior part of the magnocellular preoptic nucleus (PM), the pituitary (Pit), and the saccus vasculosus (SV). Among the EB-positive area, some cells in the PM, the anterior tuberal nucleus (NAT) and the AP were discriminated as the "blood-contacting neurons" histologically, whereas EB-positive neurons were not detected in the SD, the E, the Pit and the SV regions. In the PM, most EB-positive neurons (90 %) were immunoreactive to vasotocin (AVT) antibody, indicating that these neurons are vasotocinergic. The remaining EB-positive neurons (10 %) were not immunoreactive to ANG II and tyrosine hydroxylase (TH) antibodies. Although some neurons in the PM were immunoreactive to ANG II antibody, they were EB-negative. In contrast, almost all EB-positive neurons in the AP showed TH-like immunoreactivity (-lir), indicating that these neurons utilize catecholamine(s) as a neurotransmitter. The EB-positive neurons in the NAT were not immunoreactive to AVT, ANG II and TH antibodies, whereas some neurons without EB-staining showed ANG II-lir. Possible roles of these neurons in regulating drinking behavior in eels are discussed.

Age-associated changes in the monoaminergic innervation of rat lumbosacral spinal cord

The effects of ageing on the innervation patterns of lumbosacral spinal nuclei involved in controlling lower urinary tract functions, including micturition, were studied using immunohistochemistry for serotonin (5-HT) and tyrosine hydroxylase (TH) in male Wistar rats of 3 and 24 months. Quantitative image analysis revealed significant age-associated declines in the innervation of most regions including the intermediolateral cell nucleus, sacral parasympathetic nucleus, dorsal grey commissure and in the ventral horn including the dorsolateral nucleus which in the rat is one of the component nuclei homologous to Onuf’s nucleus in man. Notable exceptions to this generalised decline were observed in the 5-HT innervation of the sacral parasympathetic nucleus, which was maintained, and in the region of the dorsolateral motor nucleus where TH-like immunoreactivity did not significantly decline. These results suggest that the changes in micturition characteristics observed in aged rats may in part be a consequence of the alterations in, and decline of, aminergic inputs to both autonomic and somatic spinal nuclei associated with bladder function.

Mouse islets cultured with vasoactive intestinal polypeptide: effects on insulin release and immunoreactivity for tyrosine hydroxylase.

Mouse islets cultured for 1 or 4 days with or without 10 nM vasoactive intestinal polypeptide (VIP) were stained for tyrosine hydroxylase (TH) and examined for insulin secretion during culture and in a postculture perifusion system. Exposure to exogenous VIP for 4 days increased the frequency of islet cells expressing TH-like immunoreactivity. Regardless of the culturing conditions, the islets exhibited significant insulin secretory responses to 16.7 mM glucose, the effect being potentiated by 10 nM VIP in the perifusion medium. The insulin-releasing action of glucose and the potentiating effect of VIP were less pronounced in islets cultured for 1 day with VIP than in islets cultured without this neuropeptide. The following conclusions are suggested: (a) VIP stimulates the expression of TH in mouse islet cells; (b) the latency of the VIP-induced TH is a postreceptor phenomenon; (c) islet cultures exposed to VIP represent a new instance of the association between increased functional demands on beta cells and enhanced expression of TH and a new instance of VIP having trophic effects.

Noradrenergic neurons expressing substance P receptor (NK1) in the locus coeruleus complex: a double immunofluorescence study in the rat

By using a double immunofluorescence method we examined the distribution of noradrenergic neurons expressing substance P receptor (NK1) or neuromedin K receptor (NK3) in the rat brainstem. The distribution of SPR-like immunoreactive (-LI) neurons completely overlapped that of tyrosine hydroxylase (TH)-LI neurons in the locus coeruleus (A6), ventrolateral and lateral reticular formation of pons (A5 and A7). Partially overlapping distribution of SPR- and TH-LI neurons were found in certain regions of the medulla oblongata (A1–A4). Neurons showing both SPR- and TH-like immunoreactivities, however, were only found in the locus coeruleus complex (A5–A7): 100% of these TH-LI neurons displayed SPR-like immunoreactivity. Neurons showing both NKR- and TH-like immunoreactivities were not detected in the aforementioned areas of brainstem. The present study has provided morphological evidence for direct physiological modulation of noradrenergic neurons by tachykinins through SPR in locus coeruleus complex (A5–A7).

Prolactin-releasing peptide-immunoreactivity in A1 and A2 noradrenergic neurons of the rat medulla

Distribution of prolactin-releasing peptide-like immunoreactivity (PrRP-LI) was investigated in the rat medulla with the use of a rabbit polyclonal antiserum against the human PrRP-31 peptide. PrRP-positive neurons were noted mainly in two areas of the caudal medulla: ventrolateral reticular formation and commissural nucleus of the nucleus of the solitary tract (NTS), corresponding to the A1 and A2 areas. PrRP-LI neurons were absent in the medulla rostral to the area postrema. Double-labeling the sections with PrRP antisera and tyrosine hydroxylase (TH) monoclonal antibodies revealed extensive colocalization of PrRP- and TH-like immunoreactivity (TH-LI) in neurons of the A1 and A2 areas. Our results show that PrRP-LI is expressed in a population of A1 and A2 noradrenergic neurons of the rat caudal medulla.

Two groups of TH-like immunoreactive neurons in the frog ( Rana pipiens) retina

The morphology and distribution of TH-like immunoreactive (TH-IR) cells in the retina of Rana pipiens were studied in retinal whole mounts and in radial and horizontal sections. A large majority (96%) of the immunoreactive cells were found in the inner nuclear layer while a few cells were found in the ganglion cell layer. All TH-IR cells had round to oval somata with average diameter of 10 μm. The 2–4 primary processes of these cells distributed extensively to sublamina 1 of the inner plexiform layer (IPL) and sparsely to sublamina 5. Two groups of TH-IR cells were distinguished: one, designated thin cells, had only thin (<2 μm diameter) primary processes; the second, designated thick cells, had one or more primary processes with diameter(s) exceeding 2 μm for a distance of 5 μm or more from the soma. The thin cells did not significantly differ from the thick cells in soma diameter, number of primary processes, horizontal spread of processes or vertical lamination of processes. Nearest neighbor analyses of the two types revealed that the population of TH-IR cells ( thick and thin together) have an orderly distribution while the thick cells alone are more randomly distributed, indicating that the thick cells do not comprise a functional population. The total number of TH-IR cells varied between retinas; the variability was due principally to variation of thin cell density. It is hypothesized that the thick cells are a subpopulation of the TH-IR cells which are in a particular physiological state at the time of fixation.

Morphological and Functional Roles of Macrophages and Postganglionic Sympathetic Fibers in Dorsal Root Ganglions of a Rat Neuropathic Pain Model.

The induction and functional involvement of macrophages and postganglionic sympathetic fibers have been reported in rat neuropathic pain models. In the present study, we examined the morphology and structural relationship among macrophages, postganglionic sympathetic fibers, the dorsal root ganglion (DRG) neurons, and the satellite cells by light and electron microscopic examination. We used the rat spinal nerve ligation model reported by Kim & Chung. On postoperative day (POD) 1, when sympathetically maintained pain was confirmed, macrophages invaded from the nerve ligation site and reached the DRG neuronal area. Macrophages might have induced the demyelination of the spinal nerve and the degeneration of the satellite cells, and then the DRG neurons, from which the satellite cells were detached, lost the insulation from other neurons or nerve fibers. After the invasion of macrophages, postganglionic sympathetic fibers with TH-like immunoreactivity migrated into the same area, and occupied sites around the neuronal soma which were surrounded by satellite cells under normal conditions. At POD 180, the sympathetically maintained pain had disappeared, and the DRG neurons and spinal nerve showed normal structure without any invasive macrophages or postganglionic sympathetic fibers. The above changes in the morphological arrangements of macrophages and postganglionic sympathetic fibers in the DRG were thus elucidated and we suggest that this process may be part of the mechanisms underlying the development of the sympathetically maintained pain.

Expression of tyrosine hydroxylase, calcitonin gene-related peptide, substance P and protein gene product 9.5 in mouse islets transplanted under the kidney capsule

Pancreatic islets transplanted to the kidney of syngeneic mice were stained for calcitonin gene-related peptide (CGRP), substance P (SP), tyrosine hydroxylase (TH), acetylcholinesterase and the pan-neuronal marker, protein gene product 9.5 (PGP). Nerve fibers expressing TH-like immunoreactivity (TH-LI) and CGRP-LI were rare for 4 days but increased 2 (CGRP) or 6 (TH) weeks after transplantation. In 1-year-old grafts the CGRP-LI innervation resembled that in situ, while TH-LI and PGP-LI innervations were increased. SP-LI fibers remained rare throughout. Perikarya intrinsic to the islets did not show CGRP-LI or SP-LI. The results indicate a progressive ingrowth of sensory fibers into the grafts and that the TH-LI innervation becomes even more pronounced than in the pancreas. The post-transplantation reaction of islet intrinsic neurons does not involve CGRP and SP, contrasting with previous observations for vasoactive intestinal polypeptide.

The innervation of the synovium of the knee joint in the guinea pig: an immunohistochemical and ultrastructural study.

The innervation of the knee joint synovial membrane of the guinea pig, i.e., the synoviocyte layer, the subjacent connective tissue and the connective tissue region beneath, was analyzed with immunohistofluorescence and electron microscopy. A screening of the innervation with antibodies against the general axon marker -- protein gene product (PGP) 9,5 -- revealed the presence of nerve fibers distributed in various regions of the knee joint synovial membrane. Confirming previous studies, some of these nerve fibers stained with antibodies to tyrosine hydroxylase (TH), neuropeptide Y (NPY), substance P (SP), calcitonin gene-related peptide (CGRP), and vasoactive intestinal polypeptide (VIP). In addition, dynorphin (DYN)-containing fibers were detected, which have not been reported previously in normal joints. In general, the immunoreactive fibers were observed close to the synoviocytes and at blood vessels. Fibers with colocalization of NPY- and TH-like immunoreactivities (LIs), as well as of DYN- and TH-LIs were demonstrated. In the electron microscope, bundles of unmyelinated fibers as well as single fibers were found in the connective tissue region below the synoviocytes. Varicose parts of the nerve fibers contained mainly small, clear vesicles. Small and large dense-cored vesicles were also seen, but less frequently. Denser portions of the plasma membranes of some axons were observed in these regions, facing the extracellular space. Myelinated fibers were also observed in some nerve bundles. These findings emphasize the complex innervation of the synovial membrane, with nerve fibers containing a host of neuroactive substances. Altogether, these fibers are probably involved in many functions such as vasoregulation and control of synovial secretion in addition to being a source of mediators in joint inflammation.

Mesencephalic dopaminergic neurons expressing neuromedin K receptor (NK 3): a double immunocytochemical study in the rat

By using a double immunocytochemical method we examined the distribution of dopaminergic neurons expressing neuromedin K receptor (NKR; NK 3) in the rat brain. The distribution of NKR-like immunoreactive (-LI) neurons completely overlapped that of tyrosine hydroxylase (TH)-LI neurons in the retrorubral field (A8), substantia nigra (A9), ventral tegmental area and nucleus raphe linealis (A10). Completely or partially overlapping distributions of NKR- and TH-LI neurons were found in certain regions of the hypothalamus (A11–A15) and olfactory bulb (A16). Neurons showing both NKR- and TH-like immunoreactivities, however, were only found in A8–A10: All of the NKR-LI neurons displayed TH-like immunoreactivity, and about 71–86% of the TH-LI neurons expressed NKR-like immunoreactivity. The present results provided morphological evidence for physiological modulation of dopaminergic neurons by tachykinins through NKR in A8–A10.

Transiently catecholaminergic cells in the fetal rat express mRNA for the glutamate NMDAR 1 receptor

The N-methyl- d-aspartate (NMDA) subtype of the glutamate receptor has been shown to be vital to the development of the central nervous system. The purpose of this study was to determine if the neural crest-derived precursors which migrate to the primitive gut contain mRNA encoding for the NMDA receptor. Many of these enteric precursors briefly elaborate tyrosine hydroxylase (TH) and have been termed transiently catecholaminergic (TC) cells. TH-like immunoreactivity (TH-ir) serves as a marker for them. Immunocytochemistry combined with NMDAR, in situ hybridization revealed that TH-ir cells in Day 14 rat embryos do express mRNA coding for the NMDAR 1 receptor. However, the TC cells did not contain detectable levels of immunoreactivity for the NMDAR 1 receptor peptide. The absence of detectable NMDAR 1-like immunoreactivity might reflect some form of transcriptional or translational regulation, such that the onset of functional receptor activity is delayed until differentiation and/or synaptogenesis commence. Whether TC cell migration is glutamate-mediated remains unclear, since some of them successfully reached the gut without expressing NMDAR 1 message. Characterizing TC cell NMDA receptor activity and determining exactly when it is ensues will be of paramount importance to defining the role(s) of this receptor in ENS development. In conclusion, the expression of NMDAR 1 mRNA by TH-ir cells suggests a possible developmental role for this receptor.

Immunohistochemical localization of bioactive peptides and amines associated with the chromaffin tissue of five species of fish.

Biogenic peptides and amines associated with the chromaffin tissue in Atlantic cod (Gadus morhua), rainbow trout (Oncorhynchus mykiss), European eel (Anguilla anguilla), spiny dogfish (Squalus acanthias) and Atlantic hagfish (Myxine glutinosa) were identified utilizing immunohistochemical techniques. Within the posterior cardinal vein (PCV) in cod, trout and eel, a subpopulation of chromaffin cells displayed immunoreactivity to tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (D beta H) but not to phenylethanolamine-N-methyltransferase (PNMT). TH-like immunoreactivity was observed within cells in hagfish hearts. Nerve fibres displaying vasoactive intestinal peptide (VIP) immunoreactivity and pituitary adenylyl cyclase activating peptide (PACAP) immunoreactivity innervated cod, trout and eel chromaffin cells. In eel, neuropeptide Y (NPY)-like and peptide YY (PYY)-like immunoreactivity was located within cells in the PCV, including chromaffin cells. Serotonin-like immunoreactivity was observed within eel and cod chromaffin cells and in hagfish hearts. In the dogfish axillary bodies, nerves displaying TH-like, VIP-like, PACAP-like, substance P-like and galanin-like immunoreactivity were observed. These results are compared with those of other vertebrates, and potential roles for these substances in the control of catecholamine release are suggested.

Development of the catecholamine neurons in human embryos and fetuses, with special emphasis on the innervation of the cerebral cortex.

The cathecholaminergic (CA) systems have been described as appearing early in the development of the mammalian central nervous system (CNS), but their exact distribution in humans has been studied only following gestational week (g.w.) 13. Furthermore, it is not known when CA fibers initially penetrate the developing cerebral cortex. In this study, the CA cells groups and fibers are described in the human central nervous system from 6 to 13 g.w. as revealed with immunocytochemical techniques, with antibodies raised against three synthetic enzymes of the catecholamine (CA) pathway: tyrosine hydroxylase (TH), dopamine-beta-hydroxylase (DBH), and phenylethanolamine-N-methyltransferase (PNMT). At 6 g.w., TH-like immunoreactive (TH-IR) cell groups were widespread through the caudorostral extension of the CNS corresponding to the different dopaminergic mesencephalic and hypothalamic groups. Noradrenergic groups also were labeled in the medulla oblongata and in the locus coeruleus as well as in other areas in the pons. Additional TH-IR cell groups might represent a transient developmental expression of TH similar to that observed in the rat. DBH immunoreactivity labeled primarily the noradrenergic pontic cell groups and, to a lesser extent, groups located in the medulla oblongata. Rare PNMT-IR neurons were detected in the medulla oblongata only at 13 g.w. The main CA bundles described in the adult were also observed in human embryos and fetuses. At 6 g.w., TH-IR pathways extended caudorostrally within the central tegmental tract and the dorsal tegmental bundle, the latter merging with the dopaminergic mesotelencephalic pathway giving rise to the medial forebrain bundle in the basal forebrain. At 7-8 g.w., TH-IR fibers extended to the basal ganglia and the telencephalic wall. The first TH-IR and, to a much lesser extent, DBH-IR fibers penetrated the frontal lateral cortical anlage through the intermediate zone and sparsely through the marginal zone but not through the thin cortical plate. A second stream entered the telencephalic anlage frontomedially, ventral to the septal area. At 11 g.w., numerous TH-IR fibers invaded the subplate layer, but they penetrated the cortical plate only at 13 g.w. At that time, TH-IR and DBH-IR fibers had reached the occipital cortex in a rostrocaudal gradient. The appearance of well-organized CA system already in embryonic stages in humans could be of great importance for normal shaping of the nervous system as well as for development of cortical circuitry.

Immunocytochemical evidence of well-developed dopaminergic and noradrenergic innervations in the frontal cerebral cortex of human fetuses at midgestation.

The catecholaminergic (CA) innervation of the frontal lobe was visualized in 20- to 24-week-old human fetuses with immunocytochemical techniques, by use of antibodies raised against three synthetic enzymes of the CA pathway, tyrosine-hydroxylase (TH), dopamine-beta-hydroxylase (DBH), and phenylethanolamine-N-methyltransferase (PNMT). DBH-like immunoreactivity (IR) was probably labeling the noradrenergic (NA) fibers and terminals in the cerebral cortex since no PNMT-IR fibers were detected. In double-labeling TH-DBH experiments, 92-95% of the DBH-IR afferents were not labeled with TH antibodies, indicating that TH-like immunoreactivity (TH-IR) was found primarily in dopaminergic (DA) fibers. Although cortical layering had not yet occurred at this stage, the widespread CA innervation observed in the different areas and layers of the fetal frontal cortex was comparable to that previously described in the adult (Gaspar, Berger, Febvret, Vigny, and Henry: J. Comp. Neurol. 279:249-271, '89). At midgestation, the distribution of CA innervation was region and laminar specific: 1) The densest dopaminergic innervation in the cerebral cortex was located caudal to the genu of the corpus callosum: TH-IR fibers were abundant throughout all layers, from the medial telencephalon (future cingulate) to the dorsal areas (presumed motor cortices) and the lateral insular areas; 2) TH-IR fibers were less dense in the rostral prefrontal cortical anlage; 3) DBH-IR noradrenergic afferents were less numerous than the dopaminergic ones in all the cortical areas studied; 4) in all areas, the highest amount of TH and DBH-IR terminals was found in the upper subplate and in the lower part of the cortical plate, followed by the molecular layer and the intermediate zone. The deep subplate exhibited a lower number of positive fibers but contained TH-IR cell bodies. The presence of dense CA innervation in the immature cortical anlage of the human frontal lobe does not exclude a reorganization of DA and NA innervations within the different cortical layers and areas during the protracted pre- and postnatal period of development.

Effect of cervical vagotomy on catecholaminergic neurons in the cranial division of the parasympathetic nervous system

This study provides evidence of catecholaminergic neurons in the cranial division of the parasympathetic nervous system. Presumptive catecholaminergic preganglionic neurons in the dorsal motor nucleus of the vagus (DMX) were revealed by a clearcut depletion of intracellular catecholamine-synthesizing enzyme immunoreactivity induced by unilateral cervical vagotomy and identified on tissues immunocytochemically processed for tyrosine hydroxylase (TH), dopamine β-hydroxylase (DβH) or phenylethanolamine N-methyltransferase (PNMT). This experimental design was essential because of the recent failure in two species to reproduce data previously obtained in double-label (combined immunocytochemical-retrograde transport) studies. Vagotomy data confirmed three spatially-segregated populations of catecholaminergic visceromotor neurons in the DMX. These cell bodies were morphologically identical to preganglionic neurons observed on alternate tissues stained for Nissl substance or immunostained for choline acetyltransferase (ChAT), the enzyme biosynthesizing acetylcholine. Neurons in the central and medial DMX demonstrated fall-off of TH-like immunoreactivity (LI) ipsilateral to the vagotomy at levels caudal to the obex. This cell group is assumed to be predominantly dopaminergic since relatively few neurons at this level of the DMX expressed DβH-LI and none were immunostained for PNMT. A second population of immunoreactive neurons, concentrated in the rostral-lateral region of the DMX, was depleted of DβH-LI on the ipsilateral side but did not express PNMT. These visceromotor neurons may, therefore, biosynthesize noradrenaline and belong to the rostral pole of the A2 area. A third population of presumptive adrenergic vagal dorsomotor neurons in the rostral-medial DMX was depleted of TH-, DβH- and PNMT-LI at levels of the ipsilateral nucleus anterior to obex. Patterns of depletion of cytoplasmic enzyme-immunoreaction product were identical in all cases irrespective of the site of the transection or the postoperative survival period. Quantitative analysis demonstrated statistically significant loss of immunolabeled neurons in rostral and caudal subgroups of the DMX on the side ipsilateral to the vagotomy. It is concluded that catecholaminergic processes in the vagus nerve, as previously identified by the aldehyde-induced histofluorescence method, may partly arise from the lower brainstem.