M. Frances Davies

Two distinct effects of 5-hydroxytryptamine on single cortical neurons.

The ability of the indoleamine serotonin (5-hydroxytryptamine; 5-HT) to alter membrane characteristics of neocortical neurons was analyzed using intracellular recording techniques. The present study demonstrates that 5-HT primarily depolarized 68% of cortical neurons probably by decreasing a resting K+ conductance, an effect blocked by the antagonists ritanserin and cinanserin and apparently mediated by 5-HT2 receptors. A hyperpolarization associated with an increased conductance state and insensitive to 5-HT2 antagonists was observed in 26% of the neurons and could be mimicked by the selective 5-HT1A agonist (+/-)-8-hydroxy-2-(di-N-propyl-amino)tetralin (8-OH-DPAT). Therefore cortical pyramidal neurons contain at least two distinct functional 5-HT receptors whose activation produces opposing effects on membrane potential and conductance.

Isoflurane and Nociception: Spinal α2A Adrenoceptors Mediate Antinociception while Supraspinal α1 Adrenoceptors Mediate Pronociception

Background The authors recently established that the analgesic actions of the inhalation anesthetic nitrous oxide were mediated by noradrenergic bulbospinal neurons and spinal &agr;2B adrenoceptors. They now determined whether noradrenergic brainstem nuclei and descending spinal pathways are responsible for the antinociceptive actions of the inhalation anesthetic isoflurane, and which &agr; adrenoceptors mediate this effect. Methods After selective lesioning of noradrenergic nuclei by intracerebroventricular application of the mitochondrial toxin saporin coupled to the antibody directed against dopamine &bgr; hydroxylase (D&bgr;H-saporin), the antinociceptive action of isoflurane was determined. Antagonists for the &agr;1 and &agr;2 adrenoceptors were injected at spinal and supraspinal sites in intact and spinally transected rats to identify the noradrenergic pathways mediating isoflurane antinociception. Null mice for each of the three &agr;2-adrenoceptor subtypes (&agr;2A, &agr;2B, and &agr;2C) and their wild-type cohorts were tested for their antinociceptive response to isoflurane. Results Both D&bgr;H-saporin treatment and chronic spinal transection enhanced the antinociceptive effects of isoflurane. The &agr;1-adrenoceptor antagonist prazosin also enhanced isoflurane antinociception at a supraspinal site of action. The &agr;2-adrenoceptor antagonist yohimbine inhibited isoflurane antinociception, and this effect was mediated by spinal &agr;2 adrenoceptors. Null mice for the &agr;2A-adrenoceptor subtype showed a reduced antinociceptive response to isoflurane. Conclusions The authors suggest that, at clinically effective concentrations, isoflurane can modulate nociception via three different mechanisms: (1) a pronociceptive effect requiring descending spinal pathways, brainstem noradrenergic nuclei, and supraspinal &agr;1 adrenoceptors; (2) an antinociceptive effect requiring descending noradrenergic neurons and spinal &agr;2A adrenoceptors; and (3) an antinociceptive effect mediated within the spinal cord for which no role for adrenergic mechanism has been found.

A substance P receptor (NK1) antagonist can reverse vascular and nociceptive abnormalities in a rat model of complex regional pain syndrome type II.

&NA; Sciatic nerve section in rats evokes chronic hindlimb edema, pain behavior, and hyperalgesia, a syndrome resembling complex regional pain syndrome (CRPS II) in man. Furthermore, there is an increase in spontaneous protein extravasation in the hindpaw skin of rats after sciatic transection, similar to the increased protein extravasation observed in the edematous limbs of CRPS patients. Now we demonstrate that sciatic nerve section also generates chronic hindlimb warmth, distal articular tenderness, allodynia, and periarticular osteoporosis, sequelae of nerve injury resembling those observed in CRPS. We postulated that facilitated substance P signaling may contribute to these vascular and nociceptive abnormalities and attempted to reverse these changes with the long acting substance P receptor (NK1) antagonist LY303870. Hindpaw spontaneous extravasation was inhibited by LY303870. Systemic administration of LY303870 also reversed hindpaw edema and cutaneous warmth. Intrathecal, but not systemic administration of LY303870 reversed soft tissue and articular mechanical hyperalgesia in the hindpaw. Collectively, these data further support the hypothesis that the sciatic nerve transection model closely resembles CRPS and that substance P contributes to the spontaneous extravasation, edema, warmth, and mechanical hyperalgesia observed in this model.

Comparative analgesic and mental effects of increasing plasma concentrations of dexmedetomidine and alfentanil in humans.

Background: In animals, systemic and intrathecal administration of the &agr;2 -adrenergic receptor agonist dexmedetomidine results in robust antinociceptive effects in models of heat pain. In humans, systemically administered dexmedetomidine is approved for sedating patients in the intensive care unit. However, whether systemic administration of dexmedetomidine in humans produces significant analgesia at doses causing sedation but not unconsciousness remains controversial. Methods: This study in human volunteers used a placebo-controlled, double-blind, and randomized design to examine whether dexmedetomidine at doses causing mild to severe sedation produces analgesia in experimental models of heat and electrical pain. Results were compared to the effects of the &mgr;-opioid receptor agonist alfentanil. A computer-controlled infusion provided four median step-up plasma concentrations of dexmedetomidine (0.09, 0.24, 0.54, and 1.23 ng/ml) and alfentanil (13.4, 33.8, 67.8, and 126.1 ng/ml). Results: Sedative and cognitive effects of dexmedetomidine were dose-dependent, resulting in a median sedation score of 95 of 100 and slowing of cognitive speed (reaction time, trail-making test) by a factor of about two at the highest plasma concentration. Dexmedetomidine did not attenuate heat or electrical pain. Alfentanil caused severe sedation (median sedation score 88 of 100) and slowed cognitive speed by a factor of approximately 1.4 at the highest plasma concentration. Alfentanil attenuated heat and electrical pain dose dependently. Conclusion: This study documents that systemic dexmedetomidine lacks analgesic efficacy for heat and electrical pain at doses causing mild to severe sedation. These results provide further evidence suggesting that systemic administration of dexmedetomidine lacks broad analgesic activity in models of acute pain at doses not rendering humans unconscious.

Opiate receptors in the periaqueductal gray mediate analgesic effect of nitrous oxide in rats

The site of action and the pathways which are activated by nitrous oxide (N2O) to produce an analgesic effect are not well defined. Experiments were designed to determine whether N2O produces analgesia by activating opiate receptors or alpha2-adrenoceptors in periaqueductal gray. The analgesic effect of N2O was determined using the tail flick response to noxious radiant heat in lightly anesthetized rats. Different antagonists were bilaterally microinjected into ventrolateral periaqueductal gray to determine whether the analgesic effect produced by N2O was reversed. The increase in the tail flick latencies produced by N2O was reversed by bilateral microinjection into the ventrolateral part of periaqueductal gray with the opiate receptor antagonist naloxone 2.5 microg/0.5 microl, but not with the alpha2-adrenoceptors antagonist yohimbine 1.5 microg/0.5 microl. These results indicate that the N2O analgesic effect is mediated by activation of opiate receptors, but not alpha2-adrenoceptors, in the periaqueductal gray. Combined with the previous experiments that the N2O analgesic effect is reversed by intrathecal injection of an alpha2-adrenoceptor antagonist but not by an opiate receptor antagonist, it seems likely that N2O causes activation of the opiate receptors in the periaqueductal gray, which in turn activate the noradrenergic descending pathways to the spinal cord to produce the analgesic effect.

The α2A adrenoceptor and the sympathetic postganglionic neuron contribute to the development of neuropathic heat hyperalgesia in mice

&NA; We have addressed the role of the sympathetic nervous system in the development and maintenance of neuropathic pain. Using a new neuropathic mouse model, we examined the development of hyperalgesia in transgenic mice lacking functional &agr;2A adrenoceptors and in sympathectomized wild‐type mice, to determine if sympathetic–sensory coupling generates hyperalgesia. The development of neuropathic heat hyperalgesia required the presence of both the &agr;2A adrenoceptor and the sympathetic postganglionic neuron (SPGN), but the development of mechanical hyperalgesia did not require either the &agr;2A adrenoceptor or the SPGN, indicating different mechanisms of sensitization. These results suggest that the development of neuropathic heat hyperalgesia, but not mechanical hyperalgesia, requires sympathetic–sensory coupling in the peripheral nervous system. Nerve injury enhanced the analgesic efficacy of the &agr;2 adrenoceptor agonist dexmedetomidine, and paradoxically also induced an analgesic response to &agr;2 adrenoceptor antagonists. The &agr;2 agonist‐evoked analgesia to mechanical stimuli was mediated by activating central &agr;2A adrenoceptors, possibly at the spinal level. The peripherally restricted &agr;2 antagonist L659,066 evoked analgesia for heat, but not for mechanical stimuli, findings which support the hypothesis that the peripheral &agr;2 adrenoceptor plays a role in both the development and the maintenance of neuropathic heat hyperalgesia. The &agr;2 antagonist‐evoked analgesia for heat stimuli was mediated by blocking peripheral and probably central &agr;2 adrenoceptors, while the analgesia for mechanical stimuli was mediated by blocking central &agr;2A adrenoceptors. Intradermal injections with an &agr;2 agonist or antagonist had no effect on nociceptive thresholds, indicating that sympathetic–sensory coupling at the level of the cutaneous nociceptor did not contribute to the maintenance of neuropathic hyperalgesia.

Activation of α2 Adrenergic Receptors Suppresses Fear Conditioning: Expression of c-Fos and Phosphorylated CREB in Mouse Amygdala

α2 adrenergic agonists such as dexmedetomidine generally suppress noradrenergic transmission and have sedative, analgesic, and antihypertensive properties. Considering the importance of the neurotransmitter norepinephrine in forming memories for fearful events, we have investigated the acute and chronic effects of dexmedetomidine on discrete cue and contextual fear conditioning in mice. When administered before training, dexmedetomidine (10–20 μg/kg, i.p.) selectively suppressed discrete cue fear conditioning without affecting contextual memory. This behavioral change was associated with a decrease in memory retrieval-induced expression of c-Fos and P-CREB in the lateral, basolateral, and central nuclei of the amygdala. Dexmedetomidines action on discrete cue memory did not occur in α2A adrenoceptor knockout (KO) mice. When dexmedetomidine was administered after training, it suppressed contextual memory, an effect that did not occur in α2A adrenoceptor KO mice. We conclude that dexmedetomidine, acting at α2A adrenoceptors, must be present during the encoding process to decrease discrete cue fear memory; however, its ability to suppress contextual memory is likely the result of blocking the consolidation process. The ability of α2 agonists to suppress fear memory may be a valuable property clinically in order to suppress the formation of memories during stressful situations.

Meperidine exerts agonist activity at the α2B-adrenoceptor subtype

Background The opioid agonist meperidine has actions, such as antishivering, that are more pronounced than those of other opioid agonists and that are not blocked with nonselective opioid antagonists. Agonists at the &agr;2 adrenoceptors, such as clonidine, are very effective antishivering drugs. Preliminary evidence also indicates that meperidine interacts with &agr;2 adrenoceptors. The authors therefore studied the ability of meperidine to bind and activate each of the &agr;2-adrenoceptor subtypes in a transfected cell system. Methods The ability of meperidine to bind to and inhibit forskolin-stimulated cyclic adenosine monophosphate formation as mediated by the three &agr;2-adrenoceptor subtypes transiently transfected into COS-7 cells has been tested. The ability of the opioid antagonist naloxone and the &agr;2-adrenoceptor antagonists yohimbine and RX821002 to block the analgesic action of meperidine in the hot-plate test was also assessed. The ability of meperidine to fit into the &agr;2B adrenoceptor was assessed using molecular modeling techniques. Results Meperidine bound to all &agr;2-adrenoceptor subtypes, with &agr;2B having the highest affinity (&agr;2B, 8.6 ± 0.3 &mgr;m; &agr;2C, 13.6 ± 1.5 &mgr;m, P < 0.05; &agr;2A, 38.6 ± 0.7 &mgr;m). Morphine was ineffective at binding to any of the receptor subtypes. Meperidine inhibited the production of forskolin-stimulated cyclic adenosine monophosphate mediated by all receptor subtypes but was most effective at the &agr;2B adrenoceptor (&agr;2B, 0.6 &mgr;m; &agr;2A, 1.3 mm; &agr;2C, 0.3 mm), reaching the same level of inhibition (approximately 70%) as achieved with the &agr;2-adrenoceptor agonist dexmedetomidine. The analgesic action of meperidine was blocked by naloxone but not by the &agr;2-adrenoceptor antagonists yohimbine and RX821002. The modeling studies demonstrated that meperidine can fit into the &agr;2B-adrenoceptor subtype. Conclusion Meperidine is a potent agonist at the &agr;2 adrenoceptors at its clinically relevant concentrations, especially at the &agr;2B-adrenoceptor subtype. Activation of the &agr;2B receptor does not contribute significantly to the analgesic action of meperidine. This raises the possibility that some of its actions, such as antishivering, are transduced by this mechanism.

Glucocorticoid inhibition of neuropathic hyperalgesia and spinal Fos expression.

Glucocorticoids are used to treat patients suffering from neuropathic pain and complex regional pain syndromes (CRPS). Previously we found that once-daily injections of the glucocorticoid methylprednisolone had no antihyperalgesic effect in the rat sciatic nerve transection model for CRPS, but on the basis of CRPS clinical data, we hypothesized that a continuous infusion of methylprednisolone might prove effective. We further postulated that the antihyperalgesic effects of glucocorticoids were mediated by the inhibition of spinal neuron hyperactivity and by the depletion of substance P or its NK(1) receptor. This study tested the effects of continuously infused methylprednisolone in sciatic nerve-transected rats. Continuous infusion of methylprednisolone (3 mg. kg(-1). d(-1) for 21 days), started after the development of neuropathic hyperalgesia, reversed both heat and mechanical hyperalgesia over 2 wk, and this effect persisted for at least 1 wk after discontinuing methylprednisolone. In addition, continuous methylprednisolone infusion partially reversed nerve injury-evoked Fos expression in the dorsal horns, suggesting that glucocorticoids can inhibit the spinal neuron hyperactivity induced by chronic sciatic nerve transection. Finally, no changes were observed in spinal substance P or NK(1) immunoreactivity after chronic methylprednisolone infusion, suggesting that the depletion of this neuropeptide or its receptor does not contribute to the antihyperalgesic actions of glucocorticoids.

Glucocorticoid inhibition of neuropathic limb edema and cutaneous neurogenic extravasation

Sciatic nerve section in rats evokes chronic limb edema, pain behavior, and hindpaw hyperalgesia, a syndrome resembling the complex regional pain syndrome type II (CRPS II or causalgia) in man. Glucocorticoids such as methylprednisolone (MP) have been used as analgesic and anti-edematous agents in patients suffering from CRPS, and interestingly these therapeutic effects appear to persist in some patients after stopping the medication. Similar to the CRPS clinical response to glucocorticoids, we now demonstrate that chronic hindpaw edema in the sciatic transection CRPS model is reversed by a continuous infusion of MP (3 mg/kg/day over 21 days), and this anti-edematous effect persists for at least 1 week after discontinuing MP. Furthermore, there is a chronic increase in spontaneous protein extravasation in the hindpaw skin of rats after sciatic transection, similar to the increased protein extravasation observed in the edematous hands of CRPS patients. A 2-week infusion of MP (3 mg/kg/day) reduced spontaneous protein extravasation in the hindpaw skin by 80%. We postulated that increased spontaneous neurogenic extravasation resulted in development of limb edema in both the animal model and the CRPS patient, and that the anti-edematous effects of MP are due to an inhibition of spontaneous extravasation. Additional experiments examined the inhibitory effects of MP infusion on electrically-evoked neurogenic extravasation in the hindpaw skin of normal rats. MP inhibition was dose- and time-dependent, with an ED(50) of 1.2 mg/kg/day for a 14-day continuous infusion of MP, and a maximum inhibitory effect requiring 17 days of MP infusion (3 mg/kg/day). MP (3 mg/kg/day for 14 days) also blocked both capsaicin- and SP-evoked neurogenic extravasation, indicating a post-junctional inhibitory effect. Our interpretation is that increased spontaneous neurogenic extravasation in this CRPS model contributed to the development and maintenance of hindpaw edema, and that chronic MP administration dose- and time-dependently blocked neurogenic extravasation at a post-junctional level, thus reversing spontaneous extravasation and limb edema in this model.