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c-fos expression in trigeminal spinal nucleus after electrical stimulation of the hypoglossal nerve in the rat.

The expression of the immediate early gene, c-fos, was used to determine the distribution of brainstem neurons activated by stimulation of the distal hypoglossal nerve (XIIn) trunk. The traditional view of the XIIn is one of purely motor function; however, stimulation of XIIn excites neurons in the trigeminal spinal nucleus. The rationale for this study was to use c-fos expression as a marker for postsynaptic activity to define the pattern of brainstem neurons excited by XIIn stimulation. It was further hypothesized that if the afferent fibers that course within XIIn supply deep lingual tissues, then c-fos expression after direct stimulation of XIIn should display a pattern similar to that seen after chemical irritant stimulation of the deep tongue muscle. In barbiturate-anesthetized male rats electrical stimulation of XIIn produced a significant increase in Fos-positive neurons in the dorsal paratrigeminal nucleus (dPa5) and laminae I-II of caudal subnucleus caudalis (Vc) and upper cervical dorsal horn. Mustard oil injection into the deep tongue muscle also produced an increase in c-fos expression in dPa5; however, the highest density of expression occurred in laminae I-II at the dorsomedial aspect of rostral Vc. Both electrical stimulation of XIIn and mustard oil stimulation of the deep tongue increased c-fos expression in the caudal ventrolateral medulla, an autonomic relay nucleus. These results suggest that one site of innervation for afferent fibers that travel within the distal trunk of XIIn is to supply the deep tongue muscle and to terminate in the dPa5. A second group of postsynaptic neurons activated only by XIIn stimulation was located in lamina I-II in caudal portions of Vc and upper cervical dorsal horn, a laminar distribution consistent with a role for XIIn afferents in sensory or autonomic aspects of lingual function.

Afferent Pathways↗

Co-localization of fixative-modified glutamate and glutaminase in neurons of the spinal trigeminal nucleus of the rat: an immunohistochemical and immunoradiochemical analysis.

The spinal trigeminal nucleus (STN) is involved in processing orofacial sensory information, including tactile, thermal and nociceptive input, and relaying this information to higher brain centers, such as the thalamus. Very little information is available regarding the major excitatory neurotransmitters of this nucleus. The amino acid glutamate has been proposed as a major excitatory neurotransmitter in the central nervous system. In the present study, a novel monoclonal antibody, specific for fixative-modified glutamate, was utilized in conjunction with polyclonal antisera against glutaminase and aspartate aminotransferase (AATase) in an attempt to identify and map the locations of possible glutamatergic neurons in the STN. Co-localization experiments were performed by radiolabeling our monoclonal antibody and using this antibody in conjunction with the polyclonal antisera against glutaminase and AATase to evaluate the possible coexistence of glutamate with glutaminase or AATase in STN neurons. In all three subnuclei of the STN, immunohistochemically labeled neuronal profiles were observed with both of the polyclonal antisera and with the monoclonal antibody. Subnucleus caudalis contained the greatest number of labeled profiles per coronal section followed by subnucleus interpolaris and subnucleus oralis. The number and the distribution of immunoreactive profiles observed after the use of the glutaminase antiserum was comparable to that obtained with the monoclonal antibody. Co-localization experiments demonstrated that all glutaminase-like immunoreactive neurons also contained fixative-modified glutamate-like immunoradioactivity. These results suggest that glutamatergic neurons are present in the spinal trigeminal nucleus. The AATase antiserum labeled more neuronal profiles in each of the three subnuclei than did the glutaminase antiserum or the monoclonal antibody. In addition, co-localization experiments indicated that glutamate-like immunoreactivity was present in only two-thirds of AATase-like immunoreactive neuronal profiles. These findings suggest that glutaminase may be a more reliable marker of glutamatergic function than AATase.

Animals↗

Excitatory amino release within spinal trigeminal nucleus after mustard oil injection into the temporomandibular joint region of the rat.

Inflammation of the temporomandibular joint (TMJ) region evokes pain and hyperalgesia as well as causing persistent changes in the response properties of central trigeminal neurons. To determine if excitatory amino acids have a role in TMJ-induced responses, extracellular concentrations were measured in microdialysate samples from probes positioned in the spinal trigeminal nucleus (Vsp) near the transition region between subnucleus interpolaris and subnucleus caudalis (Vi/Vc) in chloralose-anesthetized rats. Injection of the selective small fiber excitant, mustard oil (20 microliters, 20% solution), into the ipsilateral TMJ region caused a transient (by 10 min) increase in glutamate (from 0.48 +/- 0.16 to 1.94 +/- 0.78 microM, P < 0.005) and aspartate (from 0.29 +/- 0.11 to 1.78 +/- 0.82 microM, P < 0.025) among sites located at the ventrolateral pole of the Vi/Vc transition region (n = 6). Samples from probes located within the ventral Vsp, but outside this Vi/Vc transition region (n = 9), did not show significant changes in amino acid concentrations. Glutamate and aspartate also increased after mustard oil injections into the contralateral TMJ region. Dialysate concentrations of serine and taurine did not change significantly after mustard oil injections. Addition of high potassium (150 mM) to the perfusate solution caused increases in glutamate and aspartate regardless of probe location. The transient and selective release of glutamate and aspartate within the Vi/Vc transition after acute irritation of the TMJ region is consistent with a proposed role for excitatory amino acids in mediating noxious sensory input from deep orofacial structures. Together with previous reports of c-fos expression, these results suggest that neurons within the ventrolateral portion of the Vi/Vc transition may serve as a relay site for the integration of sensory or reflex responses to acute inflammation of the TMJ region.

Animals↗

Stimulus-function, wind-up and modulation by diffuse noxious inhibitory controls of responses of convergent neurons of the spinal trigeminal nucleus oralis.

Extracellular unitary recordings were made from 53 spinal trigeminal nucleus oralis (Sp5O) convergent neurons in halothane-anaesthetized rats. The neurons had an ipsilateral receptive field including mainly oral or perioral regions. They responded to percutaneous electrical stimulation with two peaks of activation. The first had a short latency (4.3 +/- 0.3 ms) and low threshold (0.35 +/- 0.04 mA), whereas the second had a longer latency (68.1 +/- 3.4 ms) and higher threshold (7.3 +/- 0.5 mA). Intracutaneous injection of capsaicin (0.1%) produced a strong and rapid reduction of the long-latency responses of Sp5O convergent neurons with little effect on the short-latency responses. In most cases (73%), the long-latency responses exhibited a wind-up phenomenon during repetitive (0.66 Hz) suprathreshold electrical stimulation. These results suggest that C-fibres mediate the long-latency response of Sp5O convergent neurons. Regarding the C-fibre-evoked responses, a linear relationship between the intensity of the applied current and the magnitude of the response was found within the one to three times threshold range. The Sp5O convergent neurons also encoded the intensity of mechanical stimuli applied to the skin or mucosa in the 5-50 g ranges. The evoked activity of Sp5O convergent neurons could be suppressed by noxious heat applied to the tail (52 degrees C) and long-lasting poststimulus effects followed this. These findings show that convergent neurons in the Sp5O resemble those in the deep laminae of the spinal dorsal horn and spinal trigeminal nucleus caudalis, and further support that the Sp5O plays a part in the processing of nociceptive information from the orofacial region.

Action Potentials↗

Projections from the spinal trigeminal nucleus to the entire length of the spinal cord in the rat.

Using retrograde transport of horseradish peroxidase, projections were found from the spinal trigeminal nucleus to all levels of the spinal cord in rat. In addition to the previously described projections from nucleus caudalis, axons originate from two areas - one overlapping the nucleus caudalis and interpolaris and one in the ventral division of nucleus oralis - and descend as far as the lumbosacral cord. Projections from the nucleus caudalis and interpolaris are primarily ipsilateral to all levels of the spinal cord. Cells in the nucleus oralis innervating the cervical cord are distributed bilaterally. Projections from the nucleus oralis to the thoracic and lumbosacral segments are entirely crossed, lighter and less extensive than those to cervical levels. Trigeminospinal cells originate from several loci that strongly overlap the distributions of both trigeminotectal and trigeminocerebellar neurons. These findings thus reveal a new and more complexly organized system of trigeminospinal projections than previously recognized.

Animals↗

Nitric oxide synthase inhibition lowers activity of neurons with meningeal input in the rat spinal trigeminal nucleus.

Nitric oxide is thought to control transmitter release and neuronal activity in the spinal dorsal horn and the spinal trigeminal nucleus, where nociceptive information from extra- and intracranial tissues is processed. Extracellular impulse activity was recorded from neurons in the rat spinal trigeminal nucleus with afferent input from the cranial dura mater. In contrast to the inactive isomer D-NAME, infusion of the nitric oxide synthase inhibitor L-NAME (20 mg/kg) significantly reduced neuronal activity and increased systemic blood pressure. It is concluded that nitric oxide production contributes to the ongoing activity of sensitized neurons in the spinal trigeminal nucleus. The results suggest that nitric oxide may be involved in the generation and maintenance of primary headaches such as migraine.

Action Potentials↗

Increased expression of gamma-aminobutyric acid transporters GAT-1 and GAT-3 in the spinal trigeminal nucleus after facial carrageenan injections.

The present study aimed to elucidate the distribution of gamma-aminobutyric acid (GABA) transporters in the spinal trigeminal nucleus after carrageenan injections. Dense GAT-1 and GAT-3 but very little GAT-2 immunoreactivity was observed in the normal rat spinal trigeminal nucleus. The GAT-1-positive glial cells in the normal rat spinal trigeminal nucleus contained dense bundles of glial filaments and had features of astrocytes. Some GAT-3-positive cells contained dense bundles of glial filaments and had features of astrocytes, whilst others lacked glial filaments, and contained dense marginated heterochromatin, and had features of oligodendrocyte precursor cells. An increase in immunoreactivity to both transporters was observed on the injected but not the contralateral side 3 days after facial carrageenan injections. In rats given three further weekly injections of carrageenan and killed 3 days after the fourth injection, further increases in GAT-1 and GAT-3 immunoreactivities were observed. Electron microscopy showed that transporter immunoreactivity in the spinal trigeminal nucleus of carrageenan-injected rats was predominantly present in glial processes, showing that the increase in the number of processes observed at light microscopy was due to increased immunoreactivity in glial processes. An increased expression of GABA transporters in the carrageenan-injected spinal trigeminal nucleus could therefore result in a faster removal of GABA from the synaptic cleft of GABAergic axon terminals compared to normal rats. This could result in reduced inhibition/increased activity of the trigeminothalamic neurons in the spinal trigeminal nucleus, and could contribute to hyperalgesia after carrageenan injections.

Animals↗

Inhibitory effects of nucleus raphe magnus on neuronal responses in the spinal trigeminal nucleus to nociceptive compared with non-nociceptive inputs.

The effect of stimulation in nucleus raphe magnus on neurones in the spinal trigeminal nucleus was studied in decerebrate cats. Short trains of raphe stimulation reduced or abolished the responses of neurones in nucleus oralis and nucleus caudalis to nociceptor stimulation (tooth pulp or cornea) but had a much weaker or insignificant effect on responses to non-noxious inputs (hair movement or low intensity stimulation of the infraorbital nerve). Maximum inhibition was seen 10--50 msec following the raphe stimulation and persisted up to 300 msec. The depression of neuronal responses was paralleled by an increase in the threshold of the jaw-opening reflex evoked by tooth pulp stimulation. The relation to stimulus-produced analgesia is discussed.

Animals↗

Substance P receptor-like immunoreactive neurons in the caudal spinal trigeminal nucleus send axons to the gelatinosus thalamic nucleus in the rat.

By means of substance P receptor (SPR) immunofluorescence histochemistry combined with Fluoro-Gold (FG) fluorescent retrograde labeling, SPR-like immunoreactive neurons in the caudal spinal trigeminal nucleus of the rat were observed to send their axons to the gelatinosus thalamic nucleus with a clear ipsilateral dominance. FG/SPR double-labeled neurons were distributed mainly in the ventral part of lamina I at the rostral level of the caudal spinal trigeminal nucleus. The percentages of FG/SPR-LI neurons in the total number of SPR-LI neurons and FG-labeled neurons are 10.5% and 31.1%, respectively. The present results suggest that trigemino-gelatinosus thalamic projection neurons with SPR-LI in the caudal spinal trigeminal nucleus might receive SP-containing, nociceptive primary afferent fibers from the orofacial region and transmit nociception to the gelatinosus thalamic nucleus.

Animals↗

The corticotrigeminal projection in the cat. A study of the organization of cortical projections to the spinal trigeminal nucleus.

The projection from the cerebral cortex to the spinal trigeminal nucleus has been studied light microscopically in adult cats. Both orthograde degeneration and orthograde intra-axonal labeling techniques have been applied. Our results indicate that the projection from the coronal gyrus (face area of primary somatosensory cortex) to the spinal trigeminal complex is somatotopically organized. In subnucleus caudalis this somatotopy is organized dorsoventrally and appears to match the somatotopic distribution of the divisional trigeminal afferents. Hence cortical fibers originating from the posterior coronal gyrus (upper representation) project ventrolaterally into caudalis where division I trigeminal afferents terminate. Likewise cortical fibers from the anterior coronal gyrus (jaw and tongue representation) terminate dorsomedially in caudalis to overlap with division III trigeminal afferents. In contrast, the distribution of corticofugal afferents to the rostral spinal trigeminal subnuclei (pars interpolaris and oralis) is organized mediolaterally. Therefore in these subnuclei the cortical projection does not appear to overlap the dorsoventral lamination of the divisional trigeminal afferents. In addition, our results suggest that the cortical projection to subnucleus caudalis includes fibers which terminate in the marginal zone (lamina I) and its extensions into the spinal trigeminal tract (the interstitial cells of Cajal). We have been unable to document a projection from the proreate gyrus to the spinal trigeminal complex.

Animals↗

Both oral and caudal parts of the spinal trigeminal nucleus project to the somatosensory thalamus in the rat.

Recent evidence has been accumulated that not only spinal trigeminal nucleus caudalis (Sp5C) neurons but also spinal trigeminal nucleus oralis (Sp5O) neurons respond to noxious stimuli. It is unknown, however, whether Sp5O neurons project to supratrigeminal structures implicated in the sensory processing of orofacial nociceptive information. This study used retrograde tracing with Fluorogold in rats to investigate and compare the projections from the Sp5O and Sp5C to two major thalamic nuclei that relay ascending somatosensory information to the primary somatic sensory cortex: the ventroposteromedial thalamic nucleus (VPM) and the posterior thalamic nuclear group (Po). Results not only confirmed the existence of contralateral projections from the Sp5C to the VPM and Po, with retrogradely labelled neurons displaying a specific distribution in laminae I, III and V, they also showed consistent and similar numbers of retrogradely labelled cell bodies in the contralateral Sp5O. In addition, a topographic distribution of VPM projections from Sp5C and Sp5O was found: neurons in the dorsomedial parts of Sp5O and Sp5C projected to the medial VPM, neurons in the ventrolateral Sp5O and Sp5C projected to the lateral VPM, and neurons in intermediate parts of Sp5O and Sp5C projected to the intermediate VPM. All together, these data suggest that not only the Sp5C, but also the Sp5O relay somatosensory orofacial information from the brainstem to the thalamus. Furthermore, trigemino-VPM pathways conserve the somatotopic distribution of primary afferents found in each subnucleus. These results thus improve our understanding of trigeminal somatosensory processing and help to direct future electrophysiological investigations.

Animals↗

CGRP in the trigeminal nucleus, spinal cord and hypothalamus: effect of gonadal steroids.

Calcitonin gene-related peptide (CGRP) contents were assayed in cervical spinal cord, trigeminal nucleus and hypothalamus throughout the estrous cycle and in male and ovariectomized rats. In the trigeminal nucleus, neither testosterone nor 17 beta-estradiol seem to affect CGRP accumulation, but progesterone seems to decrease it. In the cervical spinal cord, ovarian steroids seem to decrease CGRP while testosterone does not seem to influence it. In the hypothalamus, CGRP was only detectable in the male rat suggesting a positive effect of testosterone. It had marked circadian rhythm. In conclusion, CGRP content appears to be affected by gonadal steroids in the hypothalamus, the cervical spinal cord and the trigeminal nucleus in the rat.

Animals↗

Stimulation of the spinal trigeminal nucleus supports classical conditioning of the rabbit's nictitating membrane response.

Acquisition, extinction, and differential conditioning of the rabbit's nictitating membrane response to a tone conditioned stimulus were supported by electrical stimulation of the spinal trigeminal nucleus. Stimulation of the accessory abducens nucleus, the abducens nucleus, and the reticular formation at the level of the spinal trigeminal nucleus supported lower, transient levels of conditioning. The results are discussed in terms of stimulation of sensory inputs to the brainstem and cerebellum.

Animals↗

Effect of baclofen enantiomorphs on the spinal trigeminal nucleus and steric similarities of carbamazepine.

The seemingly structurally different drugs, baclofen and carbamazepine, have a similar neurophysiologic effect on the cat spinal trigeminal nucleus and a similar clinical effect in the amelioration of trigeminal neuralgia pain. In this investigation, we report on the enhancement of segmental inhibition by carbamazepine and l-baclofen; d-baclofen produced no effect on segmental inhibition. Doses of l-baclofen one fifth its equivalent racemic dosage produced a much greater enhancement of segmental inhibition. d-Baclofen, when given prior to l-baclofen, blocked the effect of l-baclofen on segmental inhibition and the unconditioned response at previously effective doses. Pretreatment with d-baclofen also blocked the effect of subsequent carbamazepine on segmental inhibition, but had no effect on the unconditioned response. Crystallographic evaluation of carbamazepine and the enantiomorphs of baclofen revealed a surprisingly good fit of baclofen isomers to moieties of the carbamazepine molecule. The results suggest that the baclofen enantiomorphs and carbamazepine have a common mechanism of action in the cat spinal trigeminal nucleus, and that d-baclofen, though inactive, is capable of interfering with the effect of l-baclofen and to a lesser extent with carbamazepine.

Animals↗

Ultrastructural changes in acetylcholinesterase activity in the deafferented spinal trigeminal nucleus.

Acetylcholinesterase (AChE) activity was investigated in synaptic areas of the cat spinal trigeminal nucleus (pars interpolaris and pars caudalis) ipsilateral and contralateral to complete retrogasserian rhizotomy. Vibratome sections of tissue taken from animals of 1, 3, 6, 14, and 21 days survival were examined by electron microscopy following a histochemical reaction for AChE activity employing a method based on the Karnovsky-Roots technique for demonstrating reaction product. As degeneration progressed with survival time, enzymatic activity was initially reduced in synaptic clefts of injured afferent terminals and subsequently was enhanced throughout the extracellular space, including within synaptic clefts of possibly reinnervated sites. These changes in enzymatic activity with primary deafferentation are discussed in relation to the process of reinnervation, the development of neuronal hyperactivity, and possible noncholinergic functions of AChE.

Acetylcholinesterase↗