Off-cells In Rostral Ventromedial Medulla Research Articles

The inhibiting role of periaqueductal gray metabotropic glutamate receptor subtype 8 in a rat model of central neuropathic pain

ABSTRACTThe pathophysiology of neuropathic pain is very complex. It involves several environmental and central mechanisms. In this study, we tried to assess the modulatory effect of (S)-3,4-Dicarboxyphenylglycine (DCPG), a metabotropic glutamate receptor subtype 8 (mGluR8) agonist, in a model of chronic central neuropathic pain in male rats. We used a spinal cord contusion method (T6-T8) for the induction of chronic central neuropathic pain.Male Wistar rats were randomly assigned to 5 equal groups (n = 10 per group). Clips compression injury model was used to induce chronic central neuropathic pain. Three weeks after spinal cord injury DCPG, siRNA and normal saline were administered intra-ventrolaterally to the periaqueductal gray (PAG) region. Paw withdrawal response to acetone (cold allodynia) was assessed through acetone test. In addition, the effects of DCPG on rostral ventromedial medulla (RVM) off-cells activity were evaluated with immunohistochemistry. mGluR8 expressions were also measured.We found that treatment with DCPG increased pain threshold in acetone test. In addition, immunohistochemical evaluation of RVM off-cells showed that DCPG increased the suppressive function of these cells.The results revealed that activation of mGluR8 in PAG is capable to improve pain threshold via modulation of RVM off-cells activity.Abbreviations SCI: spinal cord injury; DCPG: (S)-3,4-dicarboxyphenylglycine; PAG: periaqueductal gray; siRNA: small interfering ribonucleic acid; RVM: rostral ventromedial medulla; mGluR: metabotropic glutamate receptor

Pain-facilitating medullary neurons contribute to opioid-induced respiratory depression

Respiratory depression is a therapy-limiting side effect of opioid analgesics, yet our understanding of the brain circuits mediating this potentially lethal outcome remains incomplete. Here we studied the contribution of the rostral ventromedial medulla (RVM), a region long implicated in pain modulation and homeostatic regulation, to opioid-induced respiratory depression. Microinjection of the μ-opioid agonist DAMGO in the RVM of lightly anesthetized rats produced both analgesia and respiratory depression, showing that neurons in this region can modulate breathing. Blocking opioid action in the RVM by microinjecting the opioid antagonist naltrexone reversed the analgesic and respiratory effects of systemically administered morphine, showing that this region plays a role in both the analgesic and respiratory-depressant properties of systemically administered morphine. The distribution of neurons directly inhibited by RVM opioid microinjection was determined with a fluorescent opioid peptide, dermorphin-Alexa 594, and found to be concentrated in and around the RVM. The non-opioid analgesic improgan, like DAMGO, produced antinociception but, unlike DAMGO, stimulated breathing when microinjected into the RVM. Concurrent recording of RVM neurons during improgan microinjection showed that this agent activated RVM ON-cells, OFF-cells, and NEUTRAL-cells. Since opioids are known to activate OFF-cells but suppress ON-cell firing, the differential respiratory response to these two analgesic drugs is best explained by their opposing effects on the activity of RVM ON-cells. These findings show that pain relief can be separated pharmacologically from respiratory depression and identify RVM OFF-cells as important central targets for continued development of potent analgesics with fewer side effects.

Enhanced pronociception by amygdaloid group I metabotropic glutamate receptors in nerve-injured animals

Peripheral neuropathy has been associated with structural and functional changes of the amygdala, a key player in emotions. Here we study whether peripheral neuropathy influences pain regulation by the amygdala. For this purpose, we determined discharge rates of presumably pro- and antinociceptive pain-regulatory neurons in the rostral ventromedial medulla (RVM) following microinjection of various glutamatergic compounds into the central nucleus of the amygdala. RVM neurons were recorded in pentobarbitone-anesthetized rats with a peripheral nerve injury or sham-operation. In a separate behavioral experiment, we determined whether the influence of amygdaloid administration of a glutamatergic compound on affective pain-related behavior, as assessed by an aversive place-conditioning test, is changed by neuropathy. While glutamate or an NMDA receptor antagonist in the amygdala failed to induce marked changes in discharge rates of RVM cells, amygdaloid administration of DHPG, a group I metabotropic glutamate receptor (mGluR) agonist acting on mGluR 1 and mGluR 5, increased discharge rates of presumably pronociceptive RVM ON-cells in nerve-injured but not sham-operated animals. This pronociceptive effect of DHPG was reversed by MPEP (mGluR 5 antagonist) and CPCCOEt (mGluR 1 antagonist). CHPG, an mGluR 5 agonist, failed to influence ON-cell activity and DHPG failed to influence activity of presumably antinociceptive RVM OFF-cells. Amygdaloid administration of DHPG increased and that of CPCCOEt decreased affective pain-related behavior in nerve-injured animals. The results suggest that following nerve injury, the amygdaloid group I mGluR, particularly subtype mGluR 1, has an enhanced pronociceptive effect providing a potential mechanism for emotional enhancement of pain in peripheral neuropathy.

Periaqueductal gray neurons project to spinally projecting GABAergic neurons in the rostral ventromedial medulla

The analgesic effects of morphine are mediated, in part, by periaqueductal gray (PAG) neurons that project to the rostral ventromedial medulla (RVM). Although much of the neural circuitry within the RVM has been described, the relationship between RVM neurons and PAG input and spinal output is not known. The objective of this study was to determine whether GABAergic output neurons from the PAG target RVM reticulospinal neurons. Immunocytochemistry and confocal microscopy revealed that PAG neurons project extensively to RVM neurons projecting to the spinal cord, and two-thirds of these reticulospinal neurons appear to be GABAergic (contain GAD67 immunoreactivity). The majority (71%) of PAG fibers that contact RVM reticulospinal GAD67-immunoreactive neurons also contained GAD67 immunoreactivity. Thus, there is an inhibitory projection from PAG to inhibitory RVM reticulospinal neurons. However, there were also PAG projections to the RVM that did not contain GAD67 immunoreactivity. Additional experiments were conducted to determine whether the heterogeneity in this projection can be explained by the electrophysiological character of the RVM target neurons. PAG projections to electrophysiologically defined and juxtacellularly filled ON, OFF, and Neutral cells in the RVM were examined. Similar to the pattern reported above, both GAD67- and non-GAD67-immunoreactive PAG neurons project to RVM ON, OFF, and Neutral cells in the RVM. These inputs include a GAD67-immunoreactive projection to a GAD67-immunoreactive ON cell and non-GAD67 projections to GAD67-immunoreactive OFF cells. This pattern is consistent with PAG neurons producing antinociception by direct excitation of RVM OFF cells and inhibition of ON cells.

Tonic Endovanilloid Facilitation of Glutamate Release in Brainstem Descending Antinociceptive Pathways

Activation of transient receptor potential vanilloid-1 (TRPV1) channels in the periaqueductal gray (PAG) activates OFF antinociceptive neurons of the rostral ventromedial medulla (RVM). We examined in rats the effect of intra-ventrolateral (VL)-PAG injections of TRPV1 agonists and antagonists on the nocifensive response to heat in the plantar test, neurotransmitter (glutamate and GABA) release in the RVM, and spontaneous and tail flick-related activities of RVM neurons. The localization of TRPV1 in VL-PAG and RVM neurons was examined using various markers of glutamatergic and GABAergic neurons. Intra-VL-PAG injection of capsaicin increased the threshold of thermal pain sensitivity, whereas the selective TRPV1 antagonist 5'-iodo-resiniferatoxin (I-RTX) facilitated nociceptive responses, and blocked capsaicin analgesic effect at a dose inactive per se. Intra-VL PAG capsaicin evoked a robust release of glutamate in RVM microdialysates. I-RTX, at a dose inactive per se, blocked the effect of capsaicin, and inhibited glutamate release at a higher dose. Antinociception and hyperalgesia induced by capsaicin and I-RTX, respectively, correlated with enhanced or reduced activity of RVM OFF cells. Immunohistochemical analyses suggested that several TRPV1-immunoreactive (ir) neurons in both the VL-PAG and RVM are glutamatergic and surrounded by glutamatergic and GABAergic terminals. Our data suggest that VL-PAG neurons respond to TRPV1 stimulation by releasing glutamate into the RVM, thereby activating OFF cells and producing analgesia. The results obtained with the TRPV1 antagonist alone suggest that this pathway is tonically activated by endovanilloids.

Elevation of Endocannabinoid Levels in the Ventrolateral Periaqueductal Grey through Inhibition of Fatty Acid Amide Hydrolase Affects Descending Nociceptive Pathways via Both Cannabinoid Receptor Type 1 and Transient Receptor Potential Vanilloid Type-1 Receptors

In the ventrolateral periaqueductal gray (PAG), activation of excitatory output neurons projecting monosynaptically to OFF cells in the rostral ventromedial medulla (RVM) causes antinociceptive responses and is under the control of cannabinoid receptor type-1 (CB1) and vanilloid transient receptor potential vanilloid type 1 (TRPV1) receptors. We studied in healthy rats the effect of elevation of PAG endocannabinoid [anandamide and 2-arachidonoylglycerol (2-AG)] levels produced by intra-PAG injections of the inhibitor of fatty acid amide hydrolase URB597 [cyclohexylcarbamic acid-3'-carbamoyl-biphenyl-3-yl ester] on 1) nociception in the "plantar test" and 2) spontaneous and tail-flick-related activities of RVM neurons. Depending on the dose or time elapsed since administration, URB597 (0.5-2.5 nmol/rat) either suppressed or increased thermal nociception via TRPV1 or CB1 receptors, respectively. TRPV1 or cannabinoid receptor agonists capsaicin (6 nmol) and (R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo[1,2,3,-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone mesylate [WIN55,212-2 (4 nmol)] also suppressed or enhanced nociception, respectively. URB597 dose dependently enhanced PAG anandamide and 2-AG levels, with probable subsequent activation of TRPV1/CB1 receptors and only CB1 receptors, respectively. The TRPV1-mediated antinociception and CB1-mediated nociception caused by URB597 correlated with enhanced or reduced activity of RVM OFF cells, suggesting that these effects occur via stimulation or inhibition of excitatory PAG output neurons, respectively. Accordingly, several ventrolateral PAG neurons were found by immunohistochemistry to coexpress TRPV1 and CB1 receptors. Finally, at the highest doses tested, URB597 (4 nmol/rat) and, as previously reported, WIN55,212-2 (25-100 nmol) also caused CB(1)-mediated analgesia, correlating with stimulation (possibly disinhibition) of RVM OFF cells. Thus, endocannabinoids affect the descending pathways of pain control by acting at either CB1 or TRPV1 receptors in healthy rats.

Somatodendritic morphology of on‐ and off‐cells in the rostral ventromedial medulla

The rostral ventromedial medulla (RVM) contains two classes of physiologically defined neurons, on-cells and off-cells, that are implicated in nociceptive modulation. In a continuing effort to detail the neural circuitry that underlies the activity of these two distinct neuronal types, the somatodendritic morphology of on- and off-cells was studied in the cat, rat, and ferret. In lightly anesthetized animals, on-cells increased and off-cells decreased their discharge rate during a withdrawal reflex evoked by noxious stimuli. Following their physiological characterization by using intracellular recording, on- and off-cells were injected with either horseradish peroxidase or biocytin and their somatodendritic arborizations were examined. Labeled on- and off-cells included fusiform and stellate cells of all sizes as well as large multipolar neurons. Although the somatic shape of both on- and off-cells in RVM was heterogeneous, off-cells tended to be fusiform neurons whose long axis was oriented mediolaterally. The dendritic domains of both on- and off-cells extended bilaterally past the lateral edge of the trapezoid body or pyramid and ventrally to, and sometimes including, the trapezoid body or pyramid. In contrast to their extensive mediolateral spread, the dendritic domains of both cell types were limited to the ventral half of the reticular formation and were compressed along the rostrocaudal axis. The dendritic arbor of individual on- and off-cells extended well beyond the cytoarchitectonic boundaries of any single nuclear region, within the domain delineated as the RVM. The spatial domains of the dendritic arbors of on- and off-cells are further evidence that the on- and off-cells throughout the RVM constitute an integrated unit in the modulation of nociceptive transmission.

Evidence for two classes of nociceptive modulating neurons in the periaqueductal gray

The midbrain periaqueductal gray (PAG) and the rostral ventromedial medulla (RVM) are important links in a neuronal network that modulates nociceptive transmission. In the RVM, 2 classes of cells have been identified that show changes in activity at the time of the tail-flick response (TF) elicited by noxious heat (Fields et al., 1983a). We now report that neurons in the PAG region also show changes in activity related to TF. Extracellular recordings were made from the PAG and the ventrally adjacent tegmentum at sites from which it was possible to inhibit TF using stimulating currents of 10 microA or less. Cell activity, time of TF occurrence, and tail temperature were recorded during 5 repetitions of the heat stimulus. Periresponse and peristimulus histograms were plotted with reference to the TF and tail temperature, respectively. A significant number of neurons in the PAG region showed changes in activity that preceded the TF. "Midbrain On-cells" (13.6% of the sample) displayed an abrupt increase in firing just prior to the TF. "Midbrain Off-cells" (4.4%) paused just prior to the TF. The remaining neurons (241 of 294, or 82%) did not exhibit changes in firing prior to the TF. Thus, cells with changes in activity related to the TF are present in the PAG region as well as in the RVM. The PAG has a large projection to the RVM, and microinjection of morphine in the PAG increases activity of RVM Off-cells and decreases that of RVM On-cells.(ABSTRACT TRUNCATED AT 250 WORDS)