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At least 865 records · Page 48Linked to original sources
PubMed1997

Sources of p75-nerve growth factor receptor-like immunoreactivity in the rat suprachiasmatic nucleus.

In mammals, the suprachiasmatic nucleus is critical for the generation of circadian rhythms and their entrainment to environmental cues. In the rat, the ventrolateral aspect of the suprachiasmatic nucleus receives a robust retinal input. This region also exhibits the most intense immunolabeling for the low-affinity nerve growth factor receptor in the forebrain. Our study was aimed at identifying the sources of this low-affinity nerve growth factor receptor immunoreactivity using immunohistochemistry combined with retrograde tract-tracing, and orbital enucleation. To determine the origin of the low-affinity nerve growth factor receptor immunoreactivity from sources extrinsic to the suprachiasmatic nucleus, unilateral injections of the retrograde tracer, Fluorogold, were made into the suprachiasmatic nucleus. Retrogradely labeled neurons that were also immunopositive for the low-affinity nerve growth factor receptor were found in both the basal forebrain and the retina. In the basal forebrain, such cells were found throughout its rostrocaudal extent, with the majority also being immunoreactive for the cholinergic marker, choline acetyltransferase. In the retina, cells retrogradely labeled with Fluorogold that were immunoreactive for low-affinity nerve growth factor receptor were located in the ganglion cell layer. Orbital enucleations were performed to confirm the findings observed following retrograde labeling in the retina. Unilateral orbital enucleations resulted in a significant reduction in low-affinity nerve growth factor receptor immunoreactivity in the contralateral suprachiasmatic nucleus compared to that seen on the ipsilateral side when examined one week post-surgery. Bilateral enucleations resulted in an equal decrease on both sides of the suprachiasmatic nucleus. Similar low-affinity nerve growth factor-like immunoreactivity was seen in the suprachiasmatic nucleus even two to four weeks after bilateral enucleations. Taken together, these findings suggest that low-affinity nerve growth factor receptors in the suprachiasmatic nucleus derive from multiple sources. While some receptors may be intrinsic to suprachiasmatic nucleus neurons, most appear to be of extrinsic origin and are located on axon terminals of basal forebrain cholinergic neurons and retinal ganglion cells.

Animals↗
PubMed1997

Differing influences of dopamine agonists and antagonists on Fos expression in identified populations of globus pallidus neurons.

Dopamine agonists increase the activity of globus pallidus neurons, as shown electrophysiologically and with Fos expression. More recently it has been shown that decreased D2 receptor activity also causes pallidal Fos expression. Similar responses occur in the striatum, where both dopamine agonists and D2 blockade induce Fos, although in separate neuronal populations (i.e. striatonigral and -pallidal). The present experiments investigate the possible differential dopaminergic regulation of Fos within pallidal neuronal populations, which were classified as parvalbumin-positive or -negative (with parvalbumin immunostaining), or as projecting to various target nuclei (with retrograde transport of FluoroGold iontophoresed into these nuclei). Rats with prior nigrostriatal lesions received saline, D1 agonist, or D2 agonist. Rats with no lesions received saline, combined D1/D2 agonists, or the D2 antagonist eticlopride. Two hours after drug injection, rats were perfused and their brains processed for double-labeling: either Fos staining with parvalbumin staining, or Fos or parvalbumin staining in FluoroGold-labeled sections. Overall, dopamine drug treatments induced more Fos in parvalbumin-negative than -positive cells. However, unlike dopamine agonists, eticlopride induced significant Fos only in the parvalbumin-negative cells. Dopamine agonist-induced Fos was found in pallidal neurons projecting to each of the target nuclei examined, in both normal and nigrostriatal-lesioned rats. Eticlopride-induced Fos occurred only in pallidal neurons projecting to the striatum, providing functional evidence for pallidostriatal cells without axon collaterals to other nuclei. It was also found that pallidostriatal neurons were distinguished from other projection populations by a relative lack of parvalbumin immunoreactivity. Pallidal cells respond heterogeneously to dopaminergic treatments based on their projection target and expression of parvalbumin. The smaller Fos responses in parvalbumin-containing cells may be due largely to the calcium buffering by the parvalbumin itself. Also, the pattern of Fos expression in pallidostriatal neurons suggests that dopamine regulates activity in these cells differently than in other projection populations.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1998

Calcitonin gene-related peptide increase in the rat spinal dorsal horn and dorsal column nucleus following peripheral nerve injury: up-regulation in a subpopulation of primary afferent sensory neurons.

Calcitonin gene-related peptide in sensory primary afferent neurons has an excitatory effect on postsynaptic neurons and potentiates the effect of substance P in the rat spinal dorsal horn. It has been established that calcitonin gene-related peptide expression in dorsal root ganglion neurons is depressed, and the effect of calcitonin gene-related peptide on dorsal horn neurons is attenuated, following peripheral nerve injury. We report here that a subpopulation of injured dorsal root ganglion neurons show increased expression of calcitonin gene-related peptide. Using in situ hybridization and the retrograde tracer, FluoroGold, we detected an increased number of medium- to large-sized rat dorsal root ganglion neurons projecting to the gracile nucleus that expressed alpha-calcitonin gene-related peptide messenger RNA following spinal nerve transection. Immunohistochemistry revealed a significant increase in calcitonin gene-related peptide immunoreactivity in the gracile nucleus and in laminae III-IV of the spinal dorsal horn. These results indicate that a subpopulation of dorsal root ganglion neurons express alpha-calcitonin gene-related peptide messenger RNA in response to peripheral nerve injury, and transport this peptide to the gracile nucleus and to laminae III-IV of the spinal dorsal horn. The increase of the excitatory neuropeptide, calcitonin gene-related peptide, in sites of primary afferent termination may affect the excitability of postsynaptic neurons, and have a role in neuronal plasticity following peripheral nerve injury.

Animals↗
PubMed1998

Expression of D1 receptor, D2 receptor, substance P and enkephalin messenger RNAs in the neurons projecting from the nucleus accumbens.

In situ hybridization was combined with FluoroGold retrograde labelling to determine the distribution of messenger RNAs for the D1 dopamine receptor, D2 dopamine receptor, beta-preprotachykinin or preproenkephalin in the neurons projecting from the nucleus accumbens to the ventral pallidum and the ventral tegmental area. Neurons were quantified in both the core and the shell of the nucleus accumbens to estimate the proportion of neurons projecting to the ventral pallidum or ventral tegmental area that contain transcripts for D1 receptors, D2 receptors, beta-preprotachykinin or preproenkephalin. Following the deposition of FluoroGold into the central ventral pallidum, both the core and the shell of the nucleus accumbens were retrogradely labelled, while deposits into the ventral tegmental area selectively labelled cells in the shell. A high percentage of nucleus accumbens neurons innervating the ventral tegmental area expressed messenger RNAs for D1 receptors (72%) and beta-preprotachykinin (62%), while less than 3% of the neurons contained messenger RNAs for preproenkephalin or D2 receptors. The neurons projecting to the ventral pallidum did not show the discrete distribution of transcripts as was observed in the accumbens-ventral tegmental area projection. Preproenkephalin messenger RNA was identified in 46% of the neurons innervating the ventral pallidum, and D2 receptor messenger RNA was found in approximately 40% of the cells. A large minority of neurons projecting from the nucleus accumbens to the ventral pallidum also expressed messenger RNAs for D1 receptors (37%) and beta-preprotachykinin (35%). While a higher percentage of D1 receptor, and beta-preprotachykinin messenger RNA expressing cells were located in the shell than in the core of the nucleus accumbens, the percentage tended to be higher in the core for cells expressing D2 receptors or preproenkephalin messenger RNA. These data indicate that messenger RNAs for D2 receptors and enkephalin are selectively expressed in the accumbens-pallidal projection while transcripts encoding D1 receptors and substance P are contained in the efferent projections to both the ventral pallidum and ventral tegmental area. The presence of D1 receptor and beta-preprotachykinin messenger RNAs in both mesencephalic and pallidal projections contrasts output from the striatum where the expression of D1 receptor and beta-preprotachykinin messenger RNAs is primarily restricted to the mesencephalic projection.

Animals↗
PubMed1998

Kainic acid, tetrodotoxin and light modulate expression of brain-derived neurotrophic factor in developing avian retinal ganglion cells and their tectal target.

Increasing evidence underlies the importance of neurotrophins as neuron-derived trophic signals in the developing visual system, although their precise roles are still undefined. Here we show that brain-derived neurotrophic factor messenger RNA is simultaneously expressed in a subpopulation of retinal ganglion cells and in their target during late embryogenesis. Moreover, light as well as the excitotoxin; kainic acid, induced an increase of the brain-derived neurotrophic factor messenger RNA, which could be blocked by the sodium-channel blocker; tetrodotoxin. Messenger RNA for trkB, a receptor for brain-derived neurotrophic factor, was found in the retinal ganglion cells expressing brain-derived neurotrophic factor showing that certain retinal ganglion cells express messenger RNA both for brain-derived neurotrophic factor and trkB. Furthermore, trkB messenger RNA was found in tectum, in the same layers as the brain-derived neurotrophic factor messenger RNA. These findings suggest that brain-derived neurotrophic factor expression is regulated in an activity-dependent manner during the phase of development when neuronal activity plays an important role.

Animals↗
PubMed1998

Preservation of cholinergic activity and prevention of neuron death by CEP-1347/KT-7515 following excitotoxic injury of the nucleus basalis magnocellularis.

We have identified a class of small organic molecules, derived from the indolocarbazole K-252a, that promote the survival of cultured neurons. However, many of these indolocarbazoles inhibit protein kinase C and neurotrophin-activated tyrosine kinase receptors. These kinase inhibitory activities may limit the utility of these compounds for neurological disorders. A bis-ethyl-thiomethyl analogue of K-252a, CEP-1347/KT-7515, has been identified that lacks protein kinase C and tyrosine kinase receptor inhibitory activities, yet retains the ability to promote survival of cultured neurons, including cholinergic neurons derived from the basal forebrain. In the present studies, CEP-1347/KT-7515 was assessed for neurotrophic activity on basal forebrain neurons of in vivo rats following excitotoxic insult. Ibotenate infusion into the nucleus basalis magnocellularis reduced levels of choline acetyltransferase activity in the cortex, as well as reduced numbers of choline acetyltransferase-immunoreactive and retrogradely (FluoroGold)-labelled cortically-projecting neurons in the nucleus basalis. Systemically administered CEP-1347/KT-7515 attenuated the loss of cortical choline acetyltransferase activity and the loss of the number of choline acetyltransferase-immunoreactive and retrogradely-labelled FluoroGold neurons in the nucleus basalis. Moreover, CEP-1347/KT-7515 ameliorated the loss of cortical choline acetyltransferase if administration was initiated one day, but not seven days post-lesion. Together, these results demonstrate that CEP-1347/KT-7515 protects damaged cortically-projecting basal forebrain neurons from degeneration. Thus, CEP-1347/KT-7515 may have therapeutic potential in neurodegenerative diseases, such as Alzheimer's disease, in which basal forebrain cholinergic neurons degenerate.

Animals↗
PubMed1999

Effect of graded spinal cord compression on cardiovascular neurons in the rostro-ventro-lateral medulla.

In patients with spinal cord injury, cardiovascular disturbances such as hypotension, bradycardia and autonomic dysreflexia can be directly linked to abnormalities of central autonomic control. To date, the changes in bulbospinal innervation of sympathetic preganglionic neurons after compressive spinal cord injury have not been investigated. Thus, we examined the effect of varying severity of compressive spinal cord injury on neurons of the rostro-ventro-lateral medulla, a nucleus of key importance in cardiovascular control. Adult rats with 20 g, 35 g and 50 g clip compression injuries (n= 18) of the cord at T1 and uninjured controls (n=13) were studied. Neurons in the rostro-ventro-lateral medulla with preserved spinal connections eight weeks after spinal cord injury were identified by retrograde labelling with 4% FluoroGold introduced into the cord at T6. Bulbospinal neurons in the rostro-ventro-lateral medulla were also examined immunocytochemically for the adrenaline-synthesizing enzyme phenylethanolamine-N-methyltransferase. In control rats an average of 451+/-12 rostro-ventrolateral medulla neurons were phenylethanolamine-N-methyltransferase positive. Of these, 213+/-6 projected to the T6 spinal cord. The number of rostro-ventro-lateral medulla neurons retrogradely labelled by FluoroGold decreased as a linear function of severity of spinal cord injury (r= -0.95; P<0.0001). After 50g spinal cord injury at T1, only 7+/-1 rostro-ventro-lateral medulla neurons were labelled by FluoroGold, of which 6+/-1 were phenylethanolamine-N-methyltransferase positive. Moreover, the number of phenylethanolamine-N-methyltransferase positive rostro-ventro-lateral medulla neurons decreased to 361+/-16 after 50 g spinal cord injury. We conclude that compressive spinal cord injury results in disconnection of rostro-ventro-lateral medulla neurons, which project to the thoracic spinal cord, and that these changes vary with the severity of injury. The majority of these axotomized rostro-ventro-lateral medulla neurons maintain their immunopositivity for the adrenaline-synthesizing enzyme phenylethanolamine-N-methyltransferase.

Animals↗
PubMed1999

Increased activity in the form of endurance training increases calcitonin gene-related peptide content in lumbar motoneuron cell bodies and in sciatic nerve in the rat.

The relative content of calcitonin gene-related peptide in lumbar motoneuron cell bodies (semiquantitative immunohistochemistry) and sciatic nerve was examined in rats who had previously undergone a 16-week period of endurance training on a motor-driven treadmill. Soleus motoneurons were identified in the spinal cord by their fluorescence following injection of FluoroGold into the muscle one week before killing. In sedentary rats, calcitonin gene-related peptide was detectable in 76-90% of motoneurons, with no difference in the proportions of negative cells, or in the mean staining intensity of positive cells, between soleus and neighbouring (presumptive fast hindlimb muscle) unlabelled moto-neurons. In endurance-trained rats, the estimated content of calcitonin gene-related peptide was significantly increased (90%) in cell bodies of soleus and neighbouring motoneurons, with no training-induced alterations in the proportions of calcitonin gene-related peptide-positive cells in either sample. The content of the neuropeptide was also significantly higher (37%) in sciatic nerve of endurance-trained rats. Relative accumulation of calcitonin gene-related peptide proximal to a sciatic nerve ligature applied 4 h before, however, was unchanged. The increases in calcitonin gene-related peptide in motoneuron cell bodies and sciatic nerve axons following endurance training may indicate an up-regulation of the synthesis, transport and terminal release of this neuropeptide, which could play a significant role in other morphological and functional adaptations which are known to occur at the neuromuscular junction following this chronic change in activity level.

Animals↗
PubMed1999

Differential regulation of the growth-associated proteins, GAP-43 and SCG-10, in response to unilateral cortical ablation in adult rats.

Synapse replacement after brain injury has been widely documented by anatomical studies in various parts of both the developing and adult nervous system. However, the molecular events that define the specificity of the empirically derived rules of reactive synaptogenesis in different regions of the adult brain remain unclear. In this study we examined the differential regulation of the lesion-induced response of the two growth-associated proteins, superior cervical ganglia-10 and growth-associated protein-43, after unilateral cortex ablation, and determined a hierarchical order for the lesion response from remaining afferent projection neurons originating from the contralateral cortex, ipsilateral thalamus and substantia nigra. We report that in response to unilateral cortex ablation both messenger RNA, by northern blot, and protein, by western blot, for superior cervical ganglia-10 but not growth-associated protein-43 was increased in the homologous area of the contralateral cortex but not the ipsilateral thalamus or substantia nigra. In addition, the specificity of the superior cervical ganglia-10 response, assessed by combined in situ hybridization and retrograde FluoroGold labeling of striatal afferent neurons, found that superior cervical ganglia-10 messenger RNA was increased prominently in layer V pyramidal neurons of the contralateral corticostriatal pathway but was unchanged in afferent projection neurons from the thalamus and substantia nigra. Furthermore, the increase in both superior cervical ganglia-10 messenger RNA and protein seen at three days postlesion in contralateral corticostriatal neurons coincides in time with the initiation of neurite outgrowth in the deafferented striatum by contralateral corticostriatal axons described in our previous ultrastructural study. However, if cortical input to the striatum was removed bilaterally the lesion-induced response for superior cervical ganglia-10 messenger RNA shifted secondarily to thalamostriatal neurons in the ipsilateral thalamus. These data provide evidence that superior cervical ganglia-10 and growth-associated protein-43 are differentially regulated in neurons of the contralateral corticostriatal pathway in response to unilateral cortex ablation and suggests that superior cervical ganglia-10 plays a role in the regulation of neurite outgrowth in the adult striatum after brain injury. However, the specific role that superior cervical ganglia-10 may play in reactive synaptogenesis remains unclear. In addition, our data suggest that a hierarchical order exists for the reinnervation of deafferented striatal neurons after unilateral cortex ablation with preference given to homologous axons from the contralateral cortex.

Animals↗
PubMed1999

An electrophysiological and morphological investigation of the projections of growth hormone-releasing peptide-6-responsive neurons in the rat arcuate nucleus to the median eminence and to the paraventricular nucleus.

Growth hormone-releasing peptide-6 injection induces c-fos messenger RNA expression in many arcuate nucleus neurons, and sub-populations of neurons in this region project to the hypothalamic paraventricular nucleus. We examined electrophysiologically whether arcuate nucleus neurons that project to the paraventricular nucleus also project to the median eminence, and whether these neurons are activated by systemic injection of growth hormone-releasing peptide-6. Of 116 arcuate nucleus neurons tested, 43 were antidromically-identified as projecting to the paraventricular nucleus and a further 30 as projecting to the median eminence; these populations displayed distinct electrophysiological characteristics, and contrasting patterns of orthodromic response to stimulation of the median eminence and paraventricular nucleus, indicating that these two populations are functionally distinct with limited communication between them. Only one cell was antidromically-identified as projecting to both these regions. Three of 10 arcuate nucleus neurons that projected to the paraventricular nucleus were activated by injection of growth hormone-releasing peptide-6. In parallel experiments, we examined whether Fos protein expression is induced in arcuate nucleus neurons that project to the paraventricular nucleus, as identified by retrograde-labelling with FluoroGold. Immunocytochemical studies revealed that 20% of arcuate nucleus neurons that were retrogradely-labelled from the paraventricular nucleus were Fos-positive following growth hormone-releasing peptide-6 injection, although cells that were both Fos-positive and retrogradely-labelled accounted for less than 5% of the total number of Fos-positive arcuate nucleus neurons. These results confirm that there is a direct projection from the arcuate nucleus to the paraventricular nucleus and indicate that growth hormone-releasing peptide-6 activates some of these neurons.

Animals↗
PubMed1999

Selective gastric projections of nitric oxide synthase-containing vagal brainstem neurons.

Nitric oxide has been proposed to act as an intercellular messenger in central brainstem circuits controlling gastrointestinal motility. In particular, a subpopulation of preganglionic vagal neurons of the dorsal motor nucleus of the vagus have been shown to be reduced nicotinamide adenine dinucleotide phosphate(NADPH)-diaphorase positive; NADPH-diaphorase positive preganglionic fibers are also known to make contact with enteric neurons in the stomach. No studies, however, have correlated the neurochemical phenotype of preganglionic vagal neurons to their stomach target. The purpose of this study was to identify the subpopulation of nitric oxide synthase positive vagal neurons projecting to the stomach. Fluorescent retrograde tracers were injected in the fundus, corpus or antrum (Rhodamine beads) or painted on the anterior gastric branch of the vagus (DiI); five to 15 days later the brainstem was processed for nitric oxide synthase immunoreactivity. Of the 532 DiI-labeled neurons from the vagal anterior gastric branch, 25 (4.7%, n=5 rats) were co-localized with nitric oxide synthase immunoreactivity. Of the neurons labeled following injection of rhodamine beads in the antrum (N=231 neurons, n=5 rats) or corpus (N=166 neurons, n=4 rats) only three neurons showed nitric oxide synthase immunoreactivity (two in antrum and one in corpus, respectively). Conversely, 26 of 222 neurons (12%, n=7 rats) labeled following injection of rhodamine in the fundus showed nitric oxide synthase immunoreactivity. These results provide evidence for a discrete phenotypic subpopulation of vagal motoneurons that project to the gastric fundus, and suggest that these neurons may be the ones involved in the receptive relaxation reflex.

Animals↗
PubMed1999

Serotonergic input to cholinergic neurons in the substantia innominata and nucleus basalis magnocellularis in the rat.

The aim of the present study was to determine, at the light microscopic level, whether the serotonergic fibers originating from the dorsal raphe nucleus (B7), median raphe nucleus (B8) and ventral tegmentum (B9) make putative synaptic contacts with cholinergic neurons of the nucleus basalis magnocellularis and substantia innominata. For this purpose, we utilized: (i) the anterograde transport of Phaseolus vulgaris leucoagglutinin combined with choline acetyltransferase immunohistochemistry; (ii) choline acetyltransferase/tryptophan hydroxylase double immunohistochemistry; and (iii) the FluoroGold retrograde tracer technique combined with tryptophan hydroxylase immunohistochemistry. Following iontophoretic injections of Phaseolus vulgaris leucoagglutinin in the dorsal raphe nucleus, labeling was observed primarily in the ventral aspects of the nucleus basalis magnocellularis and in the intermediate region of the substantia innominata. When Phaseolus vulgaris leucoagglutinin was combined with choline acetyltransferase immunohistochemistry, a close association between the Phaseolus vulgaris leucoagglutinin-positive fibers and cholinergic neurons was observed, even though the majority of the Phaseolus vulgaris leucoagglutinin-immunoreactive terminals seemed to establish contact with non-cholinergic elements. Following Phaseolus vulgaris leucoagglutinin injection in the median raphe nucleus, very few labeled fibers with no evident close contact with nucleus basalis magnocellularis and substantia innominata cholinergic neurons were observed. After tryptophan hydroxylase/choline acetyltransferase double immunohistochemistry, a plexus of serotonergic (tryptophan hydroxylase-positive) fibers in the vicinity of choline acetyltransferase-immunoreactive neurons of the substantia innominata and nucleus basalis magnocellularis was observed, and some serotonergic terminals have been shown to come into very close contact with the cholinergic cells. Most of the tryptophan hydroxylase-immunoreactive terminals seem to establish contacts with non-cholinergic cells. Following FluoroGold injection in the nucleus basalis magnocellularis and substantia innominata, the majority of retrogradely labeled neurons was observed mainly in the ventromedial cell group of the dorsal raphe nucleus. In this area, a minority of the FluoroGold-positive neurons was tryptophan hydroxylase immunoreactive. These findings show that serotonergic terminals, identified in very close association with the cholinergic neurons in the substantia innominata and nucleus basalis magnocellularis, derive primarily from the B7 serotonergic cell group of the dorsal raphe nucleus, and provide the neuroanatomical evidence for a direct functional interaction between these two neurotransmitter systems in the basal forebrain.

Animals↗
PubMed1999

trkA, trkB, and trkC messenger RNA expression by bulbospinal cells of the rat.

Previous research has shown that corticospinal as well as rubrospinal neurons express the high-affinity trkB and trkC receptors but not the high-affinity trkA receptor. To determine if bulbospinal neurons in other brainstem areas show the same pattern of trk receptor expression, bulbospinal cells were labelled via the injection of the retrograde tracer FluoroGold into the spinal cord. Brainstem sections were then processed for in situ hybridization using oligonucleotide probes to the trkA, trkB, and trkC receptors. The results indicated that, although trkA expression occurred in brainstem areas that contain bulbospinal neurons (e.g., the vestibular nuclei, and the pontine reticular formation), very few FluoroGold-labelled cells expressed the trkA receptor. In contrast, at least 90% of bulbospinal cells in each brainstem area examined expressed the trkB receptor. Quantitative analysis indicated differences in the level of trkB labelling between bulbospinal cells in different brainstem areas, with the highest levels seen in the locus coeruleus and magnocellular portion of the red nucleus, and the lowest levels seen in the medial and superior vestibular nuclei and the raphe obscurus. With the exception of the accessory trigeminal nucleus, over 84% of bulbospinal cells in each brainstem area also expressed the trkC receptor. TrkC receptor expression was greatest in the locus coeruleus and subcoeruleus and lowest in the accessory trigeminal nucleus, the raphe magnus, and the vestibular nuclei. Results indicate that, as with other descending pathways, virtually all bulbospinal pathways should be amenable to treatment with brain-derived neurotrophic factor, neurotrophin-4/5 or neurotrophin-3, but not nerve growth factor, following spinal cord damage.

Animals↗
PubMed1999

Characterization of the extent of pontomesencephalic cholinergic neurons' projections to the thalamus: comparison with projections to midbrain dopaminergic groups.

We sought to determine whether pontomesencephalic cholinergic neurons which we have been shown previously to project to the substantia nigra and ventral tegmental area also contribute to the thalamic activation projection from the pedunculopontine and laterodorsal tegmental nuclei. Retrograde tracing, immunohistochemical localization of choline acetyltransferase and statistical methods were used to determine the full extent of the cholinergic projection from the pedunculopontine and laterodorsal tegmental nuclei to the thalamus. Progressively larger Fluoro-Gold injections in to the thalamus proportionally labeled increasing numbers of pontomesencephalic cholinergic cells both ipsi- and contralaterally in the pedunculopontine and laterodorsal tegmental nuclei. Multiple large thalamic injections left only a small fraction of the ipsilateral pontomesencephalic cholinergic group unlabeled. This small remainder did not correspond to the populations which project to the substantia nigra and ventral tegmental area, thereby indicating that substantia nigra- and ventral tegmental area-projecting cholinergic neurons must also project to the thalamus. We examined whether there existed any set of cholinergic neurons in the pedunculopontine and laterodorsal tegmental nuclei which did not innervate a thalamic target. The distribution of descending projections of the pedunculopontine and laterodorsal tegmental nuclei demonstrated that the unlabeled remainder cannot correspond to a purely descending group. We also show that substance P-positive cholinergic cells in the laterodorsal tegmental nucleus project to the thalamus. Further studies demonstrated that the small population of cholinergic cells left unlabeled from the thalamus were the smallest sized cholinergic cells, and included two groups of small, light-staining cholinergic cells located in the parabrachial area and central gray, adjacent to the main pedunculopontine and laterodorsal tegmental nuclei cholinergic groups. These small cells, in contrast to thalamic-projecting cholinergic cells, did not stain positively for reduced nicotinamide adenine dinucleotide phosphate-diaphorase. Taken together, these results indicated that all of the reduced nicotinamide adenine dinucleotide phosphate diaphorase-positive/choline acetyltransferase-positive neurons of the pedunculopontine/laterodorsal tegmental nuclei ascend to innervate some portion of the thalamus, in addition to the other targets they innervate. These findings indicate that the diverse physiological and behavioral effects attributed to the activity of pontomesencephalic cholinergic neurons should not be dissociated from their activating effects in the thalamus.

Animals↗
PubMed2001

Evaluation of muscle re-innervation employing pre- and post-axotomy injections of fluorescent retrograde tracers.

In experimental studies on peripheral nerve repair, the possibility to objectively compare original and post-operative innervation is of decisive importance for the selection of the proper nerve-reconstruction strategy. Herewith we report serious drawbacks encountered with the standard method of pre- and post-operative intramuscular injections of widely used retrograde neuronal tracers. Labeling of rat facial motoneurons by injection of Fast-Blue (FB; Group 1), Dil (Group 2), or Fluoro-Gold (FG; Group 3) into the whisker pad muscles was followed by transection and suture of the facial nerve. Two months later, the same rats received Dil (Group 1), FG (Group 2), and FB (Group 3) injections with the same parameters as the pre-operative injections. By quantitative evaluation of single- and double-retrogradely labeled perikarya of facial motoneurons, we tried to estimate the accuracy of re-innervation. Observations through a "UV-filter" (for FB-labeled perikarya) and a "rhodamine-filter" (for Dil-labeled perikarya) in Group 1 revealed an unexpected axotomy-triggered leakage of FB which compromised the counts. After pre-operative Dil labeling, nerve suture, and post-operative FG labeling (Group 2), Dil created an extracellular deposit in the whisker pad. Thus, the uptake of pre-operative tracer by sprouts of re-growing axons compromised counts of retrogradely labeled motoneurons. Employing the "UV-filter" in Group 3 (FG-, FB-, FG+FB-labeled perikarya), the emission of FB obscured that of FG and also compromised cell counts. The use of filter sets constructed ad hoc for detection of FG and FB rendered possible an objective comparison.

Amidines↗
PubMed1998

Propriospinal neurons in the C1-C2 spinal segments project to the L5-S1 segments of the rat spinal cord.

Physiological studies indicate that neurons in the upper cervical spinal cord have descending projections to the lumbosacral spinal cord and mediate inhibition of dorsal horn neurons activated from afferent input. In the present study, retrograde tracing techniques were used to examine the distribution of propriospinal neurons in C1-C2 spinal segments that project to lumbosacral spinal segments. Fluorogold or horseradish peroxidase were injected unilaterally or bilaterally into the L5-S1 spinal segments. After 2-4 days, rats were perfused with fixative and C1-C2 spinal segments were processed for retrograde labeling. Numerous neurons were found in the C1-C2 segments. In unilaterally and bilaterally injected rats, retrogradely labeled neurons were located on both the ipsilateral and contralateral sides. Retrogradely labeled neurons were located in the following locations: lateral cervical and spinal nuclei, nucleus proprius, ventral horn and the central gray region (area X). These studies demonstrate a descending projection from C1-C2 segments to the lower lumbar and sacral spinal cord. We hypothesize that many of these C1-C2 propriospinal neurons are important in modulating responses of spinal neurons at lower segmental levels to various peripheral stimuli.

Animals↗
PubMed1999

Nucleus ambiguus and bulbospinal ventral respiratory group neurons in the neonatal rat.

The in vitro brainstem-spinal cord preparation of the neonatal rat is an important model system for studies of the respiratory control system, yet there have not been studies to anatomically characterize respiratory neuron populations in the neonate. Fluorescent retrograde tracers were used to identify bulbospinal neurons of the ventral respiratory group and motoneurons of nucleus ambiguus in neonatal rats. Fluoro-Gold injections into the C4 ventral horn labeled bulbospinal neurons within a densely packed column within the ventrolateral intermediate reticular nucleus from the level of the pyramidal decussation to the facial nucleus. This cell column corresponded closely to the location of the ventral respiratory group of the adult rat. In particular, neurons were labeled in regions corresponding to the rostral ventral respiratory group and the Bötzinger complex. Unlike adult rats, the preBötzinger complex also contained many bulbospinal neurons. Fluoro-Gold-labeled neurons were also located in the medial reticular nuclei, raphe pallidus, and obscurus and spinal vestibular nucleus. As in adult rats, bulbospinal ventral respiratory group neurons overlapped with cervical vagal motoneurons in the external formation, and partially with those in the loose formation, but not with those in the semicompact or compact formation of nucleus ambiguus. These results indicate that the distribution of bulbospinal ventral respiratory group neurons corresponds with that observed in physiological studies of neonatal rats.

Amidines↗
PubMed2001

In vivo visualization of the cochlear nerve and nuclei with fluorescent axonal tracers.

In recent years multichannel neuroprostheses have been developed which directly stimulate the central auditory pathway. Substantially these have been used in cases of total hearing loss caused by neurofibromatosis type 2 where bilateral damage to the auditory nerve prevents more peripheral stimulation. The electrode carrier of the auditory brainstem implant (ABI) is designed to be placed on the cochlear nucleus complex residing at the lateral brainstem surface. Despite altered anatomy due to tumor growth or preceding surgery, correct electrode placement is essential to maximize the variety of pitch percept elicited during electrical stimulation with the ABI without producing side-effects. In order to assist intraoperative identification of the proximal auditory nerve and cochlear nuclei, the non-toxic fluorescent axonal tracers Fast Blue or Fluorogold were injected into the cochlea of rats and Java monkeys. Four to seven days after tracer application, labeling of the eighth cranial nerve, its entrance into the brainstem and the primary radiation of auditory fibers into the cochlear nucleus could be demonstrated as colored fluorescence on the living brain under appropriate ultraviolet illumination. Additional histological processing revealed groups of retrogradely labeled neuronal cell bodies in both species. Our results suggest that this method could also be used in humans in order to aid surgeons with the proper positioning of the electrode array.

Amidines↗