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PubMed1995

Morphometric analysis of ventral mesencephalic neurons retrogradely labeled with Fluoro-Gold following injections in the shell, core and rostral pole of the rat nucleus accumbens.

Morphologically distinct subsets of mesotelencephalic neurons were sought following retrograde transport of Fluoro-Gold from iontophoretic injections relatively restricted to the medial shell, core or rostral pole subterritories of the nucleus accumbens. The diameter and lengths of Fluoro-Gold immunolabeled dendrites of medial shell-projecting neurons were less than those of core and rostral pole-projecting neurons.

Animals↗
PubMed1995

A comparative study of NADPH-diaphorase in the sympathetic preganglionic neurons of the upper thoracic cord between spontaneously hypertensive rats and Wistar-Kyoto rats.

With retrograde tracing using fluorogold injection into the superior cervical ganglion and nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry, the present comparative study revealed that the retrogradely labelled neurons in n. intermediolateralis pars funicularis (ILf) and n. intermediolateralis pars principalis (ILp) of the autonomic region in the upper thoracic cord exhibited a much stronger reactivity for NADPH-diaphorase in Wistar-Kyoto (WKY) rats than those in spontaneously hypertensive rats (SHR). It was found that in ILf in WKY rats, 77.62% of the fluorogold-labelled neurons were NADPH-d positive, while in SHR, only 56.43% of the labelled neurons were NADPH-d positive. The frequency distribution of NADPH-d positive retrogradely labelled neurons was significantly reduced in ILf of the spinal cord of SHR (U-test: P < 0.01). In ILp in WKY rats, 65.25% of fluorogold-labelled neurons were NADPH-d positive in WKY rats, while in SHR, only 56.28% of the labelled neurons were NADPH-d positive. Although the difference (P > 0.05) in the frequency of NADPH-d positive neurons in ILp between the two strains of rats was not significant, the reductions in SHR seemed considerable. Examination of the preganglionic sympathetic trunk and the superior cervical ganglion between SHR and WKY rats revealed that virtually all the NADPH-d positive fibers were derived from the sympathetic preganglionic neurons. In SHR, the NADPH-d positive fibers were not as intensely stained as those of WKY rats. This preliminary results suggest that nitric oxide, as an inhibitory neurotransmitter, may be implicated in the onset of hypertension.

Animals↗
PubMed1996

Origins of nitric oxide synthase-containing nerve fibers in the rat basilar artery with reference to the fine structure of the nerve fibers.

The origins of nitric oxide synthase (NOS)-containing nerve fibers in the rat basilar artery were studied by a combination of Fluoro-Gold retrograde tracing and immunohistochemistry. After application of Fluoro-Gold onto the middle part of the basilar artery, the dye accumulated in the sphenopalatine, otic, trigeminal, superior cervical, nodose ganglia and in the spinal ganglia at level C2 and C3. Nerve cells with NOS-like immunoreactivity were detected in the above ganglia, except for the superior cervical ganglion. Neurons that showed both NOS-like immunoreactivity and Fluoro-Gold fluorescence were numerous in the sphenopalatine and otic ganglia, and less numerous in the trigeminal, nodose and spinal ganglia. Under electron microscopy, a number of unmyelinated nerve terminals with neuronal NOS-like immunoreactivity was seen in proximity to smooth muscle cells in the tunica media of the basilar artery. These findings provide morphological evidence that NOS-containing nerve fibers in the rat basilar artery have multiple origins, and suggest that the control of posterior cerebral circulation by the parasympathetic and sensory ganglia are more complex than previously considered.

Animals↗
PubMed1996

Partial coexistence of neuropeptide Y and calbindin D28k in the trigeminal ganglion following peripheral axotomy of the inferior alveolar nerve in the rat.

Immunohistochemistry was applied to examine the correlation between neuropeptide Y (NPY) and the two calcium binding proteins (CaBPs) parvalbumin (PV) and calbindin D28k (CB) in the trigeminal ganglion following peripheral axotomy of the inferior alveolar nerve (IAN) in the rat. Five days following transection and application of FluoroGold (FG) to the cut end of the IAN, approximately 14.8% (80/539) and 18.6% (90/483) of FG-labeled IAN neurons in the trigeminal ganglion showed PV-like immunoreactivity (-LI) and CB-LI, respectively. The mean +/- S.D. area of FG-labeled PV-like immunoreactive (-IR) cells (FG/PV-IR cells) and FG/CB-IR cells were 835.9 +/- 303.1 mu m2 and 712.7 +/- 246.0 mu m2, respectively. FG/PV-IR cells were significantly larger than FG/CB-IR cells. Fourteen days following peripheral axotomy of the IAN, NPY-LI appeared in the medium- to large-sized cells. Double immunostaining revealed that approximately 3.3% (52/1569) of NPY-IR cells in the axotomized trigeminal ganglion displayed PV-LI, while approximately 26.7% (371/1392) of NPY-IR cells displayed CB-LI. The mean +/- S.D. cross-sectional areas of PV-IR and CB-IR trigeminal ganglion cells displaying NPY-LI were 819.5 +/- 265.6 mu m2 and 766.5 +/- 279.7 mu m2, respectively. There were no significant differences in the cross-sectional areas either between NPY/PV-IR cells and NPY/CB-IR cells, or between FG/PV-IR cells and NPY/PV-IR cells, or between FG/CB-IR cells and NPY/CB-IR cells. The present results indicate that injury-evoked medium- to large-sized NPY neurons were a different population from large-sized PV neurons, and NPY was partly co-localized with CB.

Animals↗
PubMed1995

A retrograde double-labeling study of uni- and bilaterally projecting retinal ganglion cells that project to the superior colliculi after unilateral eye removal at birth in the albino rat.

By injecting Fluoro-Gold and Evans-Blue into the right and left superior colliculi of the normal adult albino rats, bilaterally projecting retinal ganglion cells were labeled in the ventrotemporal crescent accounting for 37.9% of all the labeled cells, whereas in 0- and 5-day unilaterally enucleated rats these were found in the lower half of the retina accounting for 64.8% and 80.6%, respectively. Furthermore, they tended to have larger somata (type I cells).

Animals↗
PubMed1996

Correlation between different types of retinal ganglion cells and their projection pattern in the albino rat.

Injecting Fluoro-Gold (FG) and Evans-Blue (EB) into the right dLGN and SC in the adult albino rat, ipsilaterally projecting double-labeled retinal ganglion cells were mainly seen in the ventrotemporal crescent. They were mainly large sized cells. The ipsilaterally projecting double-labeled cells tended to have larger somata than the single- and double-labeled cells projecting to the contralateral superior colliculus and/or dorsal nucleus of the lateral geniculate body.

Animals↗
PubMed1996

Demonstration of direct input from the retina to the lateral habenular nucleus in the albino rat.

The projection from the retina to the habenular complex was studied using fluorescent retrograde tracers in the albino rat (Wistar, Japan Clea). Following separate unilateral injections of Fluoro-Gold (FG), Fluoro-Ruby (FR), or 4-acetamido,4- isothiocyanostilbene-2,2'-disulfonic acid (SITS) into the lateral habenular nucleus (LHB), a small population of ganglion cells was labeled sporadically, predominantly those in the nasal retina contralateral to each injection site. Most of them were small cells, ranging from 9 to 16 mu m in diameter, roughly corresponding to the type III ganglion cell in the rat retina. Additionally, all of the structures previously described as regions projecting to the LHB were confirmed. Upon re-examination of previous brain sections of albino rats which had undergone monocular enucleation, degenerating retinal nerve axons and/or their terminals, stained by a modified selective silver impregnation method, were observed in the well-documented end regions of retinal afferents as well as the LHB. The degenerating retino-habenular nerve terminals were distributed sparsely and restricted mainly to the caudal part of the LHB contralateral to the side of ocular enucleation. The present experimental data provide evidence for the existence of a non-image forming retino-habenular pathway in the albino rat. We suggest that, besides serving as a point of convergence for some of the major conduction channels of the limbic and striatal systems, the LHB may play more general integrative roles, including participation in the integration of visual information.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1996

Trigeminal-parabrachial connections: possible pathway for nociception-induced cardiovascular reflex responses.

Noxious stimulation of dental nerves elicits marked changes in cardiovascular function. In order to investigate central pathways mediating reflex changes in cardiovascular activity, immunohistochemical localization of cells expressing the immediate-early gene, c-fos, was used to identify central nervous responding to noxious electrical stimulation of mandibular, incisor tooth dentin or chemical (capsaicin) stimulation of tooth pulp in the anesthetized rat. Injections of Fluoro-Gold were made in the lateral parabrachial region to identify efferent projections from the spinal trigeminal nucleus. Electrical and chemical stimulation produced similar patterns of Fos-positive neurons in the spinal trigeminal nucleus: subnuclei caudalis, interpolaris and oralis. Fos-positive neurons were most dense in laminae I and II of the dorsomedial subnucleus caudalis with fewer Fos-positive neurons located in the interpolaris and oralis subnuclei. Sham stimulation of tooth dentin and control vehicle injections into the tooth pulp resulted in either a few weakly stained or no Fos-positive neurons in the spinal trigeminal nucleus. Cell bodies double labeled with Fluro-Gold following injections into the parabrachial region and Fos-protein subsequent to electrical stimulation of incisor tooth were present in all three subnuclei of the spinal trigeminal nucleus. The largest number of Fos-positive neurons with efferent projections to the lateral parabrachial region were located in subnucleus caudalis (32.2 +/- 5.3 S.E.M.) and fewer were located in the interpolaris (0.4 +/- 0.4 S.E.M.) and oralis (19.8 +/- 3.5 S.E.M.) subnuclei. The results demonstrate that nociceptive dental input received by the three subnuclei of the spinal trigeminal nucleus, particularly the subnucleus caudalis, is relayed to the lateral parabrachial nucleus.

Animals↗
PubMed1996

Pituitary adenylate cyclase activating polypeptide-immunoreactive sensory neurons innervate rat adrenal medulla.

Rat adrenal chromaffin cells were invested by a dense network of nerve fibers immunoreactive to pituitary adenylate cyclase activating polypeptide-38 (PACAP-IR). Immunohistochemical studies demonstrated the presence of PACAP-IR in nodose and dorsal root ganglion cells, but not in neurons of the intermediolateral cell column and other autonomic nuclei of the thoracic and upper lumbar spinal cord. Somata of the T7 to T12 paravertebral ganglia were PACAP-negative. A few lightly labeled neurons were occasionally noted in the dorsal motor nucleus of the vagus. Injection of the retrograde tracer Fluorogold into the left adrenal medulla 3 days prior to sacrifice resulted in the labeling of a population of neurons in the ipsilateral spinal cord intermediolateral cell column (T1 to L1), ipsilateral and contralateral nodose ganglia and ipsilateral dorsal root ganglia from T7 to T10 inclusive. A small number of lightly labeled somata was occasionally noted in the dorsal motor nucleus of the vagus. Combined retrograde tracing and PACAP immunohistochemistry showed that a population of Fluorogold-containing nodose and dorsal root ganglion cells were also PACAP-positive. Pre-treatment of the rats with capsaicin caused a marked reduction of the PACAP-IR in the adrenal gland as well as in the superficial layers of the dorsal horn and caudal spinal trigeminal nucleus. These findings, in conjunction with the apparent absence of PACAP-IR in spinal sympathetic preganglionic neurons, sympathetic postganglionic neurons, and dorsal motor nucleus of the vagus, raise the possibility that PACAP-IR fibers observed in the adrenal medulla are primarily sensory in origin. As a corollary, catecholamine secretion from chromaffin cells may be modulated by the peptidergic sensory afferents in addition to the cholinergic sympathetic preganglionic nerve fibers.

Adrenal Medulla↗
PubMed1996

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

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

Afferent Pathways↗
PubMed1996

Central origins of substance P-like immunoreactive fibers and terminals in the spinal trigeminal caudal subnucleus in the rat.

After trigeminal rhizotomy, some substance P-like immunoreactive (SP-LI) fibers and terminals in the spinal trigeminal caudal subnucleus (Vc), specially in its superficial laminae (laminae I and II), still remained in the rat. Employing a combination of Fluoro-Gold retrograde tracing and immunofluorescence histochemical staining for SP, we found that the main central origins of these SP-LI fibers and terminals were midbrain periaqueductal gray (PAG), nucleus raphe magnus (NRM) and other raphe nuclei, and nucleus reticularis gigantocellularis pars alpha; all of them are important structures of the endogenous pain control system. The present results provided morphological evidence for PAG or NRM stimulation could inhibit neuronal activities in the Vc evoked by orofacial nociceptive stimulation and also suggested that SP might be an important neurotransmitter or neuromodulator for endogenous pain control system.

Animals↗
PubMed1996

Evidence for divergent projections to the brain noradrenergic system and the spinal parasympathetic system from Barrington's nucleus.

The present study was designed to determine whether Barrington's nucleus, which lies ventromedial to the locus coeruleus (LC) and projects to the sacral parasympathetic nucleus, is a source of afferent projections to the LC. Restricted injections of the anterograde tracer, biocytin, into Barrington's nucleus labeled varicose fibers that extended from the injection site into the LC. Consistent with this, injections of the retrograde tracers, wheatgerm agglutinin conjugated to horseradish peroxidase coupled to gold particles (WGA-Au-HRP) or fluorescein-conjugated latex beads, into the LC labeled numerous (approximately 10%) Barrington's neurons that were also retrogradely labeled by Fluoro-Gold (FG) injections in the spinal cord. Retrograde tracing from the LC combined with corticotropin-releasing hormone (CRH) immunohistochemistry revealed that at least one third of the retrogradely labeled neurons in Barrington's nucleus were CRH-immunoreactive (CRH-IR). Finally, in triple labeling studies, CRH-Barrington's neurons were consistently observed that were retrogradely labeled from both the and spinal cord. These findings implicate Barrington's nucleus as an LC afferent and a source of CRH-IR fibers in the LC. Additionally, the results suggest that some Barrington's neurons diverge to innervate both the spinal cord and the LC. This divergent innervation may serve to coregulate the sacral parasympathetic nervous system and brain noradrenergic system, thus providing a mechanism for coordinating pelvic visceral functions with forebrain activity.

Afferent Pathways↗
PubMed1988

Evidence for dual serotonergic projections to neocortex: axons from the dorsal and median raphe nuclei are differentially vulnerable to the neurotoxin p-chloroamphetamine (PCA).

Previous studies have shown that there are morphologically dissimilar serotonergic (5-HT) axon types in rat cerebral cortex which are differentially sensitive to the neurotoxic effects of certain psychotropic drugs: methylenedioxyamphetamines (MDA and MDMA) and p-chloroamphetamine (PCA) cause degeneration of fine 5-HT axon terminals in cortex, while sparing beaded axons. Moreover, a recent anterograde transport study suggests that fine and beaded 5-HT axons arise from the dorsal raphe (DR) and median raphe (MR) nuclei, respectively. These data led us to propose that the DR projection to neocortex is selectively vulnerable to the neurotoxic effects of PCA, while the MR projection is resistant; this hypothesis was tested in the present study by comparing retrograde axonal transport of the fluorescent tracer Fluoro-Gold in PCA-treated and control rats. Using this method, only axons that survive PCA treatment can take up and transport the injected label back to the cell bodies of origin, thus allowing us to determine which raphe-cortical projections remain intact after PCA. The results show that PCA administration produces a loss of fine 5-HT axon terminals in neocortex and a concomitant reduction in the number of retrogradely labeled neurons in the DR (77% decrease), when compared to controls. In contrast, beaded 5-HT axon terminals are spared and the number of labeled neurons in the MR remains unchanged after PCA. These results demonstrate that DR and MR projections to cortex are differentially vulnerable to PCA: fine axon terminals arise from neurons in the DR and are highly sensitive to the neurotoxic effects, whereas beaded axons from the MR are resistant. We therefore propose that there are two anatomically and functionally separate 5-HT projections to cortex having different (1) nuclei of origin, (2) axon morphology, (3) regional distributions, and (4) pharmacological properties. Since the mood-altering substances MDA, MDMA, and PCA act specifically upon 5-HT axon terminals from the dorsal raphe nucleus, DR neurons may be preferentially involved in the control of affective state.

Amphetamines↗
PubMed1988

The fate of prelabeled Clarke's column neurons after axotomy.

We studied Clarke's Column of the L-1 spinal cord segment of young adult female rats after first prelabeling its neurons by the intracerebellar injection of Fluoro-Gold or true blue and subsequently axotomizing the labeled cells by a complete spinal cord transection at T-9. In control rats, the number of labeled neurons at 1, 5, 10, and 20 weeks showed a progressive decrease, probably due to leakage of dye from the cells. A much greater loss of labeled neurons was found in T-9 spinal cord-transected rats than in their matched controls. At 5 weeks after transection, loss of large neurons was somewhat offset by an increase in small neurons; neuron shrinkage was a likely cause of this increase, because small, very intensely labeled neurons were found in transected rats but not in control rats. By 10 and 20 weeks post-transection, the number of all prelabeled neurons in transected rats had sharply decreased. In transected rats, but not in controls, very significant increases in labeled astroglia and microglia and other labeled small cells were found at 5 weeks. At 10 weeks, the identifiable labeled astroglia had decreased but marked increases in microglia and other labeled small cells persisted. We conclude that, following a complete T-9 spinal cord transection, axotomized Clarke's column neurons first shrink in size and then die. Labeled reactive astrocytes, which are most evident 5 weeks after injury, probably indicate phagocytosis of axotomized neurons.

Animals↗
PubMed1990

Retrograde transport of fluoro-gold in corticospinal and rubrospinal neurons 10 and 20 weeks after T-9 spinal cord transection.

Retrograde labeling with horseradish peroxidase is greatly diminished in corticospinal and rubrospinal neurons axotomized by complete T-9 spinal cord transection. We found, 10 or 20 weeks after a complete T-9 cord transection, that the number of corticospinal and rubrospinal neurons retrogradely labeled after Fluoro-Gold insertion into a new transection at T-1 did not differ from that of controls. While transection alters uptake, transport, and/or intracellular metabolism of some transportable substances, it does not affect the ability of the neurons to be retrogradely labeled with Fluoro-Gold.

Animals↗
PubMed1990

Neurogenic period of ascending tract neurons in the upper lumbar spinal cord of the rat.

Although the neurogenic period for neurons in the lumbar spinal cord has been clearly established (Days 12 through 16 of gestation), it is not known when the neurogenesis of ascending tract neurons is completed within this period. The purpose of the present study was to determine the duration of the neurogenic period for projection neurons of the ascending tracts. To label neurons undergoing mitosis during this period, tritiated thymidine was administered to fetal rats on Embryonic (E) Days E13 through E16 of gestation. Ascending tract neurons of the lumbar cord were later (Postnatal Days 40-50) labeled in each animal with a retrograde tracer, Fluoro-Gold, applied at the site of a hemisection at spinal cord segment C3. Ascending tract neurons which were undergoing mitosis in the upper lumbar cord were double labeled, i.e., labeled with both tritiated thymidine and Fluoro-Gold. On Day E13, 89-92% of the ascending tract neurons were double labeled; on Day E14, 35-37%; and on Day E15, 1-4%. Results showed, then, that some ascending tract neurons were double labeled through Day E15 and were, therefore, proliferating in the final one-third of the neurogenic period. Ascending tract neurons proliferating on Day E15 were confined to laminae III, IV, V, and X and the nucleus dorsalis. Long tract neurons in the superficial dorsal horn (laminae I and II), on the other hand, were found to have completed neurogenesis on Day E14 of gestation. Results of the present study show that spinal neurogenesis of ascending projection neurons continues throughout most of the neurogenic period and does not completely follow the well-established ventral to dorsal gradient.

Animals↗
PubMed1991

A novel assay for the in vivo study of Schwann cells.

An in vivo assay was developed for long-term analyses of Schwann cells at the single cell level. Schwann cells were isolated from sciatic nerves and labeled with the fluorescent gold label Fluoro-Gold. Cells were then transplanted into severed sciatic nerves of young adult rats. Gold-labeled Schwann cells, identified by double-labeling with S100 antibodies, were observed up to 10 mm away from the transplant site and after 90 and 120 days of survival in vivo.

Animals↗
PubMed1991

Specific outgrowth from neurons of ventral mesencephalic grafts to the catecholamine-depleted striatum of adult hosts.

Grafts of fetal ventral mesencephalon including substantia nigra have been used to correct some motor deficits produced by unilateral destruction of the dopaminergic nigrostriatal pathway in rats. Histochemical studies have shown that dopaminergic neurons within the graft send processes from the graft to the host neuropil, wherein they form synapses. The results of numerous immunocytochemical studies indicate, however, that a large proportion of neurons in grafts are not catecholaminergic. Whether or not the nondopaminergic neurons in grafts project to the host brain is unknown. The purpose of the present study was to combine immunocytochemistry and retrograde tracing with fluorogold to identify the cell types which project from grafts to the host striatum. Tissue from the ventral mesencephalon of E15 fetuses was placed into the 6-hydroxydopamine denervated striatum of graft recipients. Six weeks to 6 months following transplantation, fluorogold was pressure injected under stereotaxic control immediately adjacent to the ventral mesencephalic grafts; after 4 days CNS tissue was prepared for light microscopic immunocytochemistry. Ventral mesencephalic grafts contained cell bodies immunoreactive for enkephalin, GAD, substance P, and serotonin in addition to those immunoreactive for tyrosine hydroxylase. Some cells of each immunochemically defined type were retrogradely labeled by the fluorogold injection into the host brain. Nevertheless, more catecholaminergic and serotonergic cells projected from grafts to the fluorogold injection site than did other cell types. Since many of the nonmonoaminergic neurons in grafts are probably projection neurons, our results suggest that the extent of neurite outgrowth from grafted cells is influenced by the surrounding target tissue.

Animals↗