Tyrosine Hydroxylase-like Immunoreactivity Research Articles

A role for dopamine in the peripheral sensory processing of a gastropod mollusc.

Histological evidence points to the presence of dopamine (DA) in the cephalic sensory organs of multiple gastropod molluscs, suggesting a possible sensory role for the neurotransmitter. We investigated the sensory function of DA in the nudipleuran Pleurobranchaea californica, in which the central neural correlates of sensation and foraging behavior have been well characterized. Tyrosine hydroxylase-like immunoreactivity (THli), a signature of the dopamine synthetic pathway, was similar to that found in two other opisthobranchs and two pulmonates previously studied: 1) relatively few (<100) THli neuronal somata were observed in the central ganglia, with those observed found in locations similar to those documented in the other snails but varying in number, and 2) the vast majority of THli somata were located in the peripheral nervous system, were associated with ciliated, putative primary sensory cells, and were highly concentrated in chemotactile sensory organs, giving rise to afferent axons projecting to the central nervous system. We extended these findings by observing that applying a selective D2/D3 receptor antagonist to the chemo- and mechanosensory oral veil-tentacle complex of behaving animals significantly delayed feeding behavior in response to an appetitive stimulus. A D1 blocker had no effect. Recordings of the two major cephalic sensory nerves, the tentacle and large oral veil nerves, in a deganglionated head preparation revealed a decrease of stimulus-evoked activity in the former nerve following application of the same D2/D3 antagonist. Broadly, our results implicate DA in sensation and engender speculation regarding the foraging-based decisions the neurotransmitter may serve in the nervous system of Pleurobranchaea and, by extension, other gastropods.

Effects of sciatic nerve transection on ultrastructure, NADPH-diaphorase reaction and serotonin-, tyrosine hydroxylase-, c-Fos-, glucose transporter 1- and 3-like immunoreactivities in frog dorsal root ganglion

Frogs have been used as an alternative model to study pain mechanisms. Since we did not find any reports on the effects of sciatic nerve transection (SNT) on the ultrastructure and pattern of metabolic substances in frog dorsal root ganglion (DRG) cells, in the present study, 18 adult male frogs (Rana catesbeiana) were divided into three experimental groups: naive (frogs not subjected to surgical manipulation), sham (frogs in which all surgical procedures to expose the sciatic nerve were used except transection of the nerve), and SNT (frogs in which the sciatic nerve was exposed and transected). After 3 days, the bilateral DRG of the sciatic nerve was collected and used for transmission electron microscopy. Immunohistochemistry was used to detect reactivity for glucose transporter (Glut) types 1 and 3, tyrosine hydroxylase, serotonin and c-Fos, as well as nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase). SNT induced more mitochondria with vacuolation in neurons, satellite glial cells (SGCs) with more cytoplasmic extensions emerging from cell bodies, as well as more ribosomes, rough endoplasmic reticulum, intermediate filaments and mitochondria. c-Fos immunoreactivity was found in neuronal nuclei. More neurons and SGCs surrounded by tyrosine hydroxylase-like immunoreactivity were found. No change occurred in serotonin- and Glut1- and Glut3-like immunoreactivity. NADPH-diaphorase occurred in more neurons and SGCs. No sign of SGC proliferation was observed. Since the changes of frog DRG in response to nerve injury are similar to those of mammals, frogs should be a valid experimental model for the study of the effects of SNT, a condition that still has many unanswered questions.

Olfactory bulb recovery following reversible deafferentation with repeated detergent application in the adult zebrafish

The neuroplasticity and regenerative properties of the olfactory system make it a useful model for studying the ability of the nervous system to recover from damage. We have developed a novel method for examining the effects of long-term deafferentation and regeneration of the olfactory organ and resulting influence on the olfactory bulb in adult zebrafish. To test the hypothesis that repeated damage to the olfactory epithelium causes reduced olfactory bulb afferent input and cessation of treatment allows recovery, we chronically ablated the olfactory organ every 2-3 days for 3 weeks with the detergent Triton X-100 while another group was allowed 3 weeks of recovery following treatment. Animals receiving chronic treatment showed severe morphological disruption of the olfactory organ, although small pockets of epithelium remained. These pockets were labeled by anti-calretinin, indicating the presence of mature olfactory sensory neurons (OSNs). Following a recovery period, the epithelium was more extensive and neuronal labeling increased, with three different morphologies of sensory neurons observed. Repeated peripheral exposure to Triton X-100 also affected the olfactory bulb. Bulb volumes and anti-tyrosine hydroxylase-like immunoreactivity, which is an indicator of afferent activity, were diminished in the olfactory bulb of the chronically treated group compared to the control side. In the recovery group, there was little difference in bulb volume or antibody staining. These results suggest that repeated, long-term nasal irrigation with Triton X-100 eliminates a substantial number of mature OSNs and reduces afferent input to the olfactory bulb. It also appears that these effects are reversible and regeneration will occur in both the peripheral olfactory organ and the olfactory bulb when given time to recover following cessation of treatment. We report here a new method that allows observation not only of the effects of deafferentation on the olfactory bulb but also the effects of reinnervation.

Comparative immunohistochemistry of the cephalic sensory organs in Opisthobranchia (Mollusca, Gastropoda)

The structure and function of the central nervous systems of opisthobranch gastropods have been studied extensively. However, the organisation and function of the peripheral nervous system are poorly understood. The cephalic sensory organs (CSOs) are known to be chemosensory structures in the head region of opisthobranchs. In the present study, we used immunohistochemical methods and confocal laserscanning microscopy to comparatively examine the CSOs of different opisthobranchs, namely Acteon tornatilis, Aplysia punctata, Archidoris pseudoargus and Haminoea hydatis. We wanted to characterise sensory epithelia in order to infer the function of sensory structures and the organs they constitute. Immunoreactivity against the three antigens tyrosine hydroxylase, FMRFamide and serotonin was very similar in the CSOs of all investigated species. Tyrosine hydroxylase-like immunoreactivity was detected primarily in subepidermal sensory cell bodies, which were much more abundant in the anteriorly situated CSOs. This observation indicates that these cells and the respective organs may be involved in contact chemoreception and mechanoreception. The dominant features of FMRFamide-like immunoreactivity, especially in the posterior CSOs, were tightly knotted fibres which reveal glomerulus-like structures. This suggests an olfactory role for these organs. Serotonin-like immunoreactivity was detected in an extensive network of efferent fibres, but was not found within any peripheral cell bodies. Serotonin-like immunoreactivity was found in the same glomerulus-like structures as FMRFamide-like immunoreactivity, indicating a function of serotonin in the efferent control of olfactory inputs. Besides this functional implication, this study could also add some knowledge on the doubtful homology of the CSOs in opisthobranch gastropods.

Dopaminergic control of foregut contractions in Locusta migratoria

Tyrosine hydroxylase-like immunoreactivity is present in cell bodies and processes in the brain and optic lobes of Locusta migratoria, with processes projecting along the frontal connectives to form a neuropile within the frontal ganglion. Immunoreactive cell bodies and processes are also evident in the hypocerebral and ventricular ganglia with processes extending over the foregut. Tyrosine hydroxylase is the rate-limiting enzyme in dopamine biosynthesis, and high-performance liquid chromatography coupled to electrochemical detection was used to confirm the presence of dopamine in the innervation to the foregut. Spontaneous foregut contractions are under the control of the ventricular ganglia and are absent when these ganglia are removed. Dopamine leads to an inhibition of both the amplitude and frequency of phasic contractions of the foregut that are produced when the ventricular ganglia are left attached. Dopamine has direct effects on the foregut muscle in the absence of the ventricular ganglia, inhibiting a proctolin-induced contraction in a dose-dependent manner.

Nicotinamide adenine dinucleotide is released from sympathetic nerve terminals via a botulinum neurotoxin A-mediated mechanism in canine mesenteric artery

Using high-performance liquid chromatography techniques with fluorescence and electrochemical detection, we found that beta-nicotinamide adenine dinucleotide (beta-NAD) is released in response to electrical field stimulation (4-16 Hz, 0.3 ms, 15 V, 120 s) along with ATP and norepinephrine (NE) in the canine isolated mesenteric arteries. The release of beta-NAD increases with number of pulses/stimulation frequencies. Immunohistochemistry analysis showed dense distribution of tyrosine hydroxylase-like immunoreactivity (TH-LI) and sparse distribution of TH-LI-negative nerve processes, suggesting that these blood vessels are primarily under sympathetic nervous system control with some contribution of other (e.g., sensory) neurons. Exogenous NE (3 micromol/l), alpha,beta-methylene ATP (1 micromol/l), neuropeptide Y (NPY, 0.1 micromol/l), CGRP (0.1 micromol/l), vasoactive intestinal peptide (VIP, 0.1 micromol/l), and substance P (SP, 0.1 micromol/l) had no effect on the basal release of beta-NAD, suggesting that the overflow of beta-NAD is evoked by neither the sympathetic neurotransmitters NE, ATP, and NPY, nor the neuropeptides CGRP, VIP, and SP. Botulinum neurotoxin A (BoNTA, 0.1 micromol/l) abolished the evoked release of NE, ATP, and beta-NAD at 4 Hz, suggesting that at low levels of neural activity, release of these neurotransmitters results from N-ethylmaleimide-sensitive factor attachment protein receptor/synaptosomal-associated protein of 25 kDa-mediated exocytosis. At 16 Hz, however, the evoked release of NE, ATP, and beta-NAD was reduced by BoNTA by approximately 90, 60, and 80%, respectively, suggesting that at higher levels of neural activity, beta-NAD is likely to be released from different populations of synaptic vesicles or different populations of nerve terminals (i.e., sympathetic and sensory terminals).

Transmitter contents of cells and fibers in the cephalic sensory organs of the gastropod mollusc Phestilla sibogae.

While the central ganglia of gastropod molluscs have been studied extensively, relatively little is known about the organization and functions of the peripheral nervous system in these animals. In the present study, we used immunohistochemical procedures to examine the innervation of the rhinophores, oral tentacles and region around the mouth of the aeolid nudibranch, Phestilla sibogae. Serotonin-like immunoreactivity was found in an extensive network of efferent projections apparently originating from central neurons, but was not detected within any peripheral cell bodies. In contrast, large numbers of peripheral, and presumably sensory, somata exhibited reactivity to an antibody raised against tyrosine hydroxylase (the enzyme catalyzing the initial step in the conversion of tyrosine into the catecholamines). Additional tyrosine hydroxylase-like immunoreactivity was detected in afferent fibers of the peripheral cells and in several cells within the rhinophoral ganglia. The presence of serotonin, dopamine and norepinephrine in the rhinophores, tentacles and central ganglia was confirmed using high-performance liquid chromatography. Finally, FMRFamide-like immunoreactivity was detected in cells and tangles of fibers found within the rhinophore, possibly revealing glomerulus-like structures along olfactory pathways. FMRFamide-like immunoreactivity was also found in somata of the rhinophoral ganglia, in a small number of cells located in the body wall lateral to the tentacles and in what appeared to be varicose terminals of efferent projections to the periphery. Together, these results indicate several new features of the gastropod peripheral nervous system and suggest future experiments that will elucidate the function of the novel cells and innervation patterns described here.

The effects of ageing and of DSP-4 administration on the micturition characteristics of male Wistar rats

This study sought to determine the effects of ageing on the in vivo micturition characteristics of male Wistar rats and to assess whether they might be replicated in young rats by using the neurotoxin DSP-4 to lesion locus coeruleus-derived noradrenergic pathways projecting to spinal cord nuclei controlling micturition. Significant age-related changes in micturition patterns were observed. There was a loss of a diurnal rhythm in micturition patterns and a large increase in voided volume, maximal between 21 and 24 months, which was paralleled by an increased water intake. DSP-4 lesions neither altered micturition patterns nor water intake in the young adult rat. DSP-4 induced changes in the pattern of tyrosine hydroxylase-like immunoreactivity (TH-LI), most notably almost complete depletion of TH-LI in the dorsolateral nucleus and retention of TH-LI in lumbosacral autonomic preganglionic nuclei, did not mimic the changes in the pattern of TH-LI seen in aged rats.

Glucoprivation-induced Fos expression in the hypothalamus and medulla oblongata in female rats.

Glucoprivation induced by 2-deoxy-D-glucose (2DG) suppresses pulsatile luteinizing hormone (LH) secretion in female rats. The suppression is enhanced in the presence of estrogen. In the present study, 2DG-induced Fos expression was examined in the solitary tract nucleus (NTS), hypothalamic paraventricular nucleus (PVN), raphe obscurus nucleus (ROb) and raphe pallidus nucleus (RPa), which have been previously suggested to be involved in glucoprivation-induced suppression of LH secretion in female rats. Ovariectomized (OVX) or estrogen-primed ovariectomized (OVX+E(2)) rats were injected intravenously with 2DG (400 mg/kg BW). The brain was removed 1 h after the injection. The number of Fos-like-immunoreactive (Fos-li) cells in the PVN and NTS was significantly increased in OVX+E(2) rats compared with control groups, but did not show a significant increase in the OVX group. Few Fos-li cells were observed in the ROb and RPa in all groups. All of the Fos-li cells in the PVN and NTS were neurons because they had immunoreactivities to microtubule-associated protein 2. Some Fos-li cells (8.3%) had tyrosine hydroxylase-like immunoreactivities in the NTS in 2DG-treated OVX+E(2) rats. These results suggest that neurons in the PVN and NTS are involved in the estrogen-dependent neural cascade mediating glucoprivic suppression of LH secretion in female rats.

Catecholamine-containing cells in the central nervous system and periphery of Aplysia californica.

Previous studies have suggested the presence of numerous catecholamine-containing cells in both the central ganglia and peripheral tissues of Aplysia, but they often offered conflicting or incomplete accounts of numbers, locations, and morphologies. The current study combines aldehyde-induced histofluorescence and tyrosine hydroxylase-like immunoreactivity together with confocal microscopy to provide details of these cells. Approximately 35-50 neurones in the cerebral ganglia, 4-8 neurones in the pedal ganglia, 5 neurones in the buccal ganglia, and numerous small fibres in various nerve trunks exhibited both immunoreactivity and aldehyde-induced fluorescence. Approximately 20 cells in the pedal ganglia and 4 cells in the buccal ganglia exhibited only immunoreactivity whereas 15-20 neurons in the cerebral ganglia exhibited only aldehyde-induced fluorescence. No somata in the pleural or abdominal ganglia exhibited aldehyde-induced fluorescence or immunoreactivity. Both aldehyde-induced histofluorescence and immunoreactivity also labelled what appeared to be two classes of catecholamine-containing cells in the gill, siphon, oesophagus, rhinophore, tentacle, and reproductive organs. The more numerous, but smaller cells had subepithelial somata and processes penetrating the overlying body wall, thus suggesting a sensory function. Another class of neurones had larger somata, often located more deeply within the tissue, and occasionally appeared to be multipolar. Processes from these various peripheral cells appeared to comprise the major component of afferent fibres and to form an extensive peripheral plexus, often associated with various muscles. The morphologies of the peripheral cells thus suggest involvement in both local and centrally mediated reflexes and responses, but additional studies must test such hypothesised functions and determine the sensory modalities that the cells mediate.

Cotransmission from sympathetic vasoconstrictor neurons: differences in guinea-pig mesenteric artery and vein

Vasoconstrictor responses to electrical field stimulation (EFS, 0.2–32 Hz, 0.1 ms, 12 V, for 1 min) were measured in endothelium-denuded segments of guinea-pig mesenteric vein and compared to responses in mesenteric artery. The distribution of both tyrosine-hydroxylase-like immunoreactivity (TH-LI) and neuropeptide Y-like immunoreactivity (NPY-LI) was also studied using anti-TH and anti-NPY antibodies. The effect of exogenous NPY (10 nM) on EFS (8 Hz, 0.3 ms, 12 V, for 1 min)-evoked overflow of noradrenaline (NA) was also studied using an HPLC technique with electrochemical detection. Veins responded with contractions at lower frequencies of stimulation than arteries. Prazosin (0.1 μM) abolished the EFS-evoked contractions in artery at 0.5–32 Hz and in vein at 0.2–1 Hz of stimulation. However, in vein, the contractile responses to EFS at 2–32 Hz of stimulation were only reduced by prazosin. Phentolamine (1 μM) abolished the responses to 0.5–4 Hz and reduced the responses to 8–32 Hz of EFS in artery. In vein, phentolamine (1 μM) abolished the responses to 0.2–1 Hz and facilitated the contractions elicited by 16–32 Hz. The NPY-receptor antagonist BIBP3226 (1 μM), in combination with phentolamine, abolished contractions in vein. Yohimbine (0.1 μM) abolished the responses to lower frequencies of stimulation in both artery (0.5–2 Hz) and vein (0.2–1 Hz). The responses to greater frequency stimulation were not affected by yohimbine in artery, and were facilitated in vein. Pre-treatment of animals for 24 h with reserpine abolished contractile responses to EFS in artery, whereas in vein, responses to 0.2–2 Hz were abolished while responses to 4–32 Hz were unchanged. Suramin (100 μM) or α,β-methylene ATP (α,βMeATP; 10–100 μM) treatment did not affect the contractile responses to EFS in either artery or vein. Pyridoxal-phosphate-6-azophenyl-2′,4′-disulfonic acid tetrasodium (PPADS; 30 μM), even potentiated the responses to 2–16 Hz in vein. However, following reserpine-treatment, both PPADS and suramin reduced the nerve-evoked contractions of vein. Either BIBP3226 (1 μM) alone or BIBP3226 in combination with PPADS or suramin abolished the contractile response to EFS in reserpine-treated veins. NPY (100 nM) produced significantly more contraction in vein than in artery (i.e., 93±2.5 versus 7±4% of the response to 70 mM KCl, respectively). NPY (10 nM) significantly reduced the NA overflow evoked by EFS at 8 Hz. Flat mount preparations and cryostat sections of both mesenteric artery and vein revealed that TH-LI and NPY-LI were co-localized in a dense network of fibers within the adventitial layer. In conclusion, NA exclusively mediates the contractile response to sympathetic nerve stimulation in guinea-pig mesenteric artery, whereas at least three neurotransmitters [i.e., NA, adenosine 5′-triphosphate (ATP) and NPY] are involved in the neural response of mesenteric vein.

Neuropeptide Y is a cotransmitter with norepinephrine in guinea pig inferior mesenteric vein

Neuropeptide Y (NPY) is a cotransmitter with noradrenaline in guinea pig inferior mesenteric vein. Tyrosine hydroxylase-like immunoreactivity and NPY-like immunoreactivity were colocalized in a dense network of fibers within the adventitial layer of guinea-pig inferior mesenteric vein. Vasoconstrictor responses to electrical field stimulation (0.2–64 Hz, 0.1 ms, 12 V, for 10 s) appear to be mediated primarily by norepinephrine at 0.2 to 4 Hz and by NPY at 8 to 64 Hz. NPY Y1 receptors mediate the contractile responses to both endogenous and exogenous NPY. Norepinephrine and NPY are involved in neuromuscular transmission in guinea pig mesenteric vein suggesting that the sympathetic nervous system requires the coordinated action of norepinephrine and NPY to serve capacitance.

Selenium deficiency potentiates methamphetamine-induced nigral neuronal loss; comparison with MPTP model

The present study was designed to understand the role of an antioxidant, selenium (Se) on methamphetamine (MA)-induced dopaminergic cell damage in the substantia nigra (SN). Male C57BL/6J mice were fed either selenium-deficient (<0.01 ppm Se) or selenium-replete (0.2 ppm Se) diet for 90 days. Se-deficiency potentiates MA-induced reductions of tyrosine hydroxylase-like immunoreactivity (TH-IR), dopamine (DA) and its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanilic acid (HVA) in the SN. These dopaminergic toxicities were comparable to that induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). By contrast, Se-repletion significantly blocked dopaminergic toxicity after MA treatments. These results suggest that Se-deficient MA-treated mouse is a relevant model of Parkinsonism, and that optimal level of Se plays a crucial role in preventing nigral dopaminergic toxicity induced by MA. However, different mechanisms in the thermoregulation mediated by MA or MPTP remain to be further determined.

Haloperidol induces Fos expression in the globus pallidus and substantia nigra of cynomolgus monkeys

Systemic injections of the dopamine antagonist haloperidol (0.1–2.5 mg/kg) induced a dose dependent increase in Fos-like immunoreactivity (FLI) in the internal segment of the globus pallidus (GPi) and in the substantia nigra (SN) of cynomolgus monkeys. These findings are consistent with models of basal ganglia organization which predict that blockade of dopamine receptors should result in a disinhibition of cells in these structures. In the GPi, labeling was most pronounced along the ventral, lateral and medial borders of the nucleus and none of the pallidal cells expressing FLI were immunopositive for choline acetyltransferase. In the SN, immunoreactive nuclei were concentrated in the pars reticulata and the majority of labeled nigral neurons did not display tyrosine hydroxylase-like immunoreactivity. A small number of cells displaying FLI were also observed in the external pallidal segment, but no labeling was seen in the subthalamic nucleus. These findings indicate that blockade of dopamine receptors induces a characteristic pattern of Fos expression in the primate brain which strongly resembles that previously reported in rodents.

Serotonin-like immunoreactivity in the stomatogastric nervous systems of crayfishes from four genera.

We used whole-mount immunocytochemistry to characterize the distribution of serotonin in the stomatogastric nervous systems of seven species of crayfish representing three genera from the family Cambaridae (Orconectes, Cambarus, and Procambarus) and one from the family Astacidae (Pacifastacus). In all species, we observed serotonin-like immunoreactivity in four gastropyloric receptor (GPR) neurons located in the lateral ventricular nerves, with one pair of neurons in each nerve. As in other crustaceans, the GPR axons project to the stomatogastric ganglion and to the bilateral commissural ganglia. In three crayfishes, we observed the GPR axons crossing the commissural ganglia, and extending toward the thoracic nervous system. This feature was most clearly and consistently seen in Pacifastacus leniusculus. The number of stained somata in the commissural ganglia varied among crayfish species from two (in Procambarus clarkii) to five (in Pacifastacus leniusculus). The largest soma (the L cell) displayed both serotonin- and tyrosine hydroxylase-like immunoreactivity in all species, suggesting that serotonin and dopamine are cotransmitters in this cell. The inferior esophageal nerve and a branch of this nerve (the inner labral nerve) contained several axons with serotonin-like immunoreactivity. These axons were clearly present in only one species (Procambarus clarkii). Serotonin acts as a neuromodulator of rhythms produced by circuits in the crab and lobster stomatogastric ganglion, and is likely to play a similar role in crayfish. Differences are apparent in the distribution of serotonin among crayfish species and between crayfish and other crustaceans, and could result in differences in the physiological action of this modulator.

Development of catecholaminergic neurons in the pond snail, Lymnaea stagnalis: I. Embryonic development of dopamine-containing neurons and dopamine-dependent behaviors.

The embryonic development of the catecholaminergic system of the pond snail, Lymnaea stagnalis, was investigated by using chromatographic and histochemical methods. High performance liquid chromatography suggested that dopamine was the only catecholamine present in significant concentrations throughout the embryonic development of Lymnaea. Dopamine first became detectable at about embryonic stage (E) 15 (15% of embryonic development) and then increased in amount during early development to reach about 120-140 fmol per animal by around E40. Dopamine content remained stable during mid-embryogenesis (E40-65), increased slowing for the next couple of days, and then increased rapidly to culminate at about 400 fmol per animal by hatching. The detection of aldehyde- and glyoxylate-induced fluorescence and of tyrosine hydroxylaselike immunoreactivity indicated that the first catecholaminergic cells appeared in the late trochophore or early veliger stage of embryonic development (E32-35). The paired perikarya of these transient apical catecholaminergic (TAC) neurons were located beneath the apical plate, remained outside of the central ganglia during embryogenesis, and no longer contained detectable catecholamines close to hatching. TAC neurons bore cilia on the ends of short processes that penetrated the overlying epithelium; their long processes branched repeatedly under the ciliated apical plate. Several smaller catecholaminergic cells first appeared in the anterior margin of the foot at a stage when the embryos began to metamorphose from the veliger form (E55). Similar bipolar cells later appeared in the tentacle and lips. The axons of all of these small peripheral cells projected centrally and terminated within the neuropil of different central ganglia. Central catecholaminergic neurons, including RPeD1, differentiated only after metamorphosis was complete (E75). Development of locomotor, respiratory, and feeding behaviors correlated with maturation of catecholaminergic neurons, as indicated by histology and chromatography.

Coexistence of enkephalin- and tyrosine hydroxylase-like immunoreactivities in nerve fibers of the temporomandibular joint capsule of the guinea pig.

The innervation of joints has been suggested to play an important role for their normal function as well as for the pathogenesis of chronic arthritic conditions. It is therefore important to elucidate the functional anatomy of this innervation. The expression of methionine enkephalin-like immunoreactivity (ENK-LI) and tyrosine hydroxylase (TH)-LI as well as their possible colocalization were examined in the temporomandibular joint of the guinea pig: Nerve fibers with ENK-LI were demonstrated in the synovium of the guinea pig temporomandibular joint capsule with the use of indirect immunofluorescence. The ENK+ fibers were found close to the surface of the synovial membrane, deeper in the synovium, and at the walls of blood vessels. Coexistence of ENK- and TH-LI was observed frequently in the synovial nerve fibers. After removal of the superior cervical ganglion (SCG), the ENK-containing nerve fibers were reduced substantially in number on the operated side or were completely absent if the cervical sympathetic trunk was also excised. The findings indicate that the majority of fibers with ENK-LI derive from the SCG ENK may act as a neuromodulator on the catecholaminergic nerves and may also have an analgesic effect in the joint.

Regionally different noradrenergic innervation and receptor distribution in submucous plexus of guinea-pig intestine.

Regional differences of noradrenergic innervation and noradrenergic α2 receptor distribution in the submucous plexus of guinea-pig caecum, large intestine, and rectum were investigated with immunofluorescence confocal microscopy. In the caecal submucous plexus, three dimensional basket-like arrangements of small and bright dots of tyrosine hydroxylase-like immunoreactivity (TH-LIR) were observed, whereas noradrenergic α2A receptor-like immunoreactivity (NAR-LIR) was detected in somata. Both TH-LIR and NAR-LIR were localized in neuronal somata of the transverse colon. A few TH-LIR positive cell somata and many TH-LIR positive spots forming chains passing through ganglia and interganglionic nerve strands were observed, but no NAR-LIR were seen in the submucous plexus of the descending colon. There were a few TH-LIR positive somata and puncta, but no NAR-LIR were observed in the ascending colon and rectum. All of these observations show the distinct regional differences in noradrenergic innervation and noradrenergic α2A receptor distribution in the guinea-pig submucous plexus.

Association of postganglionic sympathetic neurons with primary afferents in sympathetic-sensory co-cultures.

Functional coupling between sympathetic postganglionic neurons and sensory neurons is thought to play an essential role in the pathogenesis of certain chronic pain syndromes following peripheral tissue and nerve injury. The mechanism(s) underlying this interaction are enigmatic. The relative anatomical inaccessibility of sympathetic and sensory neurons in vivo complicates study of their interrelationships. We have developed a system for long-term co-culturing of explants of sympathetic chain ganglia and dorsal root ganglia from newborn rats. Co-cultures were labelled for tyrosine hydroxylase-like immunoreactivity and studied at the light and electron microscopic levels. Explanted ganglia of both types survived well in co-culture. They maintained their tissue type-specific histological properties, including neuronal and glial morphology, and characteristic glial-neuronal associations. Moreover, neurons maintained their characteristic neurochemical identity, at least to the extent that sympathetic neurons continued to express tyrosine hydroxylase and dorsal root ganglion neurons did not. Sympathetic neurons emitted numerous outgrowing processes (axons) some of which came into association with sensory neurons in the explanted dorsal root ganglia. Some apparently specific sympathetic-sensory contacts were observed, suggesting that a functional interaction may develop between sympathetic axons and sensory neurons in vitro.

5-Hydroxytryptamine immunoreactivity is detectable in sympathetic nerve fibres in rat oral tissues.

The aim of this investigation was to examine if 5-hydroxytryptamine (5-HT) is detectable not only in mast cells but also in sympathetic nerve fibres in oral sites of the rat, including the periodontal ligament, pulp, palatal mucosa, and vestibular sulcus. Antibodies against 5-HT and tyrosine hydroxylase were used. Maxillae from rats were dissected free, fixed, decalcified, cut transversally, and processed for immunohistochemistry. Nerve fibres showing 5-HT-like immunoreactivity were regularly observed in the walls of the arteries and arterioles in the vestibular sulcus and the periodontal ligament. However, 5-HT-like immunoreactivity was not seen in the walls of the vessels of the palatal mucosa. Interestingly, 5-HT-like immunoreactivity coexisted with tyrosine hydroxylase-like immunoreactivity in the innervation of the periodontal ligament and the vestibular sulcus. Thus, the present study gives morphological correlate for the occurrence of effects of 5-HT derived not only from mast cells but also from sympathetic nerve fibres in oral tissues. The source of 5-HT in the nerve fibres as well as the functional implications of the observations remain to be determined.