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The role of 5-HT receptors in fluoxetine-induced nociception and antinociception in rats was assessed. Formalin produced a typical pattern of flinching and licking/lifting behaviors. Local peripheral ipsilateral, but not contralateral, pre-treatment with fluoxetine (0.3-3 nmol/paw) increased in a dose-dependent fashion 0.5% formalin-induced nociception. In contrast, intrathecal pretreatment with fluoxetine (0.3-3 nmol/rat) prevented nociception induced by formalin. The peripheral pronociceptive effect of fluoxetine was prevented by the 5-HT2A (ketanserin, 3-10 pmol/paw), 5-HT2B (3-(2-[4-(4-fluorobenzoyl)-1-piperidinyl]ethyl)-2,4(1H,3H)-quinazolinedione(+) tartrate, RS-127445, 3-10 pmol/paw), 5-HT2C (8-[5-(2,4-dimethoxy-5-(4-trifluoromethylphenylsulphonamido) phenyl-5-oxopentyl]1,3,8-triazaspiro[4.5] decane-2,4-dione hydrochloride, RS-102221, 3-10 pmol/paw), 5-HT3 (ondansetron, 3-10 nmol/paw), 5-HT4 ([1-[2-methylsulphonylamino ethyl]-4-piperidinyl]methyl 1-methyl-1H-indole-3-carboxylate, GR-113808, 3-100 fmol/paw), 5-HT6 (4-iodo-N-[4-methoxy-3-(4-methyl-1-piperazinyl)phenyl]benzene-sulfonamide hydrochloride, SB-258585, 3-10 pmol/paw) and 5-HT7 ((R)-3-(2-(2-(4-methylpiperidin-1-yl) ethyl) pyrrolidine-1-sulfonyl) phenol hydrochloride, SB-269970, 0.3-1 nmol/paw), but not by the 5-HT1A (N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinylcyclohexanecarboxamide maleate, WAY-100635, 0.3-1 nmol/paw), 5-HT1B/1D (N-[4-methoxy-3-(4-methyl-1-piperazinyl)phenyl]-2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)-1,1'-biphenyl-4-carboxamide hydrochloride hydrate, GR-127935, 0.3-1 nmol/paw), 5-HT1B (1'-methyl-5-[[2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)biphenyl-4-yl]carbonyl]-2,3,6,7-tetrahydrospiro[furo[2,3-f]indole-3,4'-piperidine hydrochloride, SB-224289, 0.3-1 nmol/paw), 5-HT1D (4-(3-chlorophenyl)-α-(diphenylmethyl)-1-piperazineethanol hydrochloride, BRL-15572, 0.3-1nmol/paw) nor 5-HT5A ((N-[2-(dimethylamino)ethyl]-N-[[4'-[[(2-phenylethyl)amino]methyl][1,1'-biphenyl]-4-yl]methyl]cyclopentanepropanamide dihydrochloride, SB-699551, 1-3 nmol/paw), receptor antagonists. In marked contrast, the spinal antinociceptive effect of fluoxetine was prevented by the 5-HT1A (WAY-100635, 0.3-1 nmol/rat), 5-HT1B/1D (GR-127935, 0.3-1 nmol/rat), 5-HT1B (SB-224289, 0.3-1 nmol/rat), 5-HT1D (BRL-15572, 0.3-1 nmol/rat) and 5-HT5A (SB-699551, 1-3 nmol/rat), but not by the 5-HT2A (ketanserin, 3-10 pmol/rat), 5-HT2B (RS-127445, 3-10 pmol/rat), 5-HT2C (RS-102221, 3-10 pmol/rat), 5-HT3 (ondansetron, 3-10 nmol/rat), 5-HT4 (GR-113808, 3-100 fmol/rat), 5-HT6 (SB-258585, 3-10 pmol/rat) nor 5-HT7 (SB-269970, 0.3-1 nmol/rat), receptor antagonists. These results suggest that fluoxetine produces nociception at the periphery by activating peripheral 5-HT2A/2B/2C/3/4/6/7 receptors. In addition, intrathecal fluoxetine produces antinociception by activation of spinal 5-HT1A/1B/1D/5A receptors., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Peripherally restricted analgesics are desirable to avoid central nervous system (CNS) side effects of opioids. Nonsteroidal anti-inflammatory drugs produce peripheral analgesia but have significant toxicity. GABA(B) receptors represent peripheral targets for analgesia but selective GABA(B) agonists like baclofen cross the blood-brain barrier. Recently, we found that the CNS-impermeant amino acid, isovaline, produces analgesia without apparent CNS effects. On observing that isovaline has GABA(B) activity in brain slices, we examined the hypothesis that isovaline produces peripheral analgesia mediated by GABA(B) receptors. We compared the peripheral analgesic and CNS effect profiles of isovaline, baclofen, and GABA (a CNS-impermeant, unselective GABA(B) agonist). All three amino acids attenuated allodynia induced by prostaglandin E2 injection into the mouse hindpaw and tested with von Frey filaments. The antiallodynic actions of isovaline, baclofen, and GABA were blocked by the GABA(B) antagonist, CGP52432, and potentiated by the GABA(B) modulator, CGP7930. We measured Behavioural Hyperactivity Scores and temperature change as indicators of GABAergic action in the CNS. ED(95) doses of isovaline and GABA produced no CNS effects while baclofen produced substantial sedation and hypothermia. In a mouse model of osteoarthritis, isovaline restored performance during forced exercise to baseline values. Immunohistochemical staining of cutaneous layers of the analgesic test site demonstrated co-localization of GABA(B1) and GABA(B2) receptor subunits on fine nerve endings and keratinocytes. Isovaline represents a new class of peripherally restricted analgesics without CNS effects, mediated by cutaneous GABA(B) receptors., (Copyright © 2012 IBRO. Published by Elsevier Ltd. All rights reserved.)

Cocaine's multiple pharmacological substrates are ubiquitously present in the peripheral and central nervous system. Thus, upon its administration, cocaine acts in the periphery before directly acting in the brain. We determined whether cocaine alters ventral tegmental area (VTA) neuronal activity via its peripheral actions. In urethane-anesthetized rats, we recorded VTA neuron's responses to intravenous injections of two cocaine analogs: cocaine-hydrochloride (HCl, 0.25 mg/kg), which readily cross the blood-brain barrier (BBB), and cocaine-methiodide (MI, 0.33 mg/kg), which does not cross the BBB. Both cocaine analogs produced sustained changes in discharge rates that began 5 s after the initiation of a 10-s drug infusion. Within the first 90 s post-injection, the magnitudes of neuronal responsiveness of both cocaine analogs were comparable, but later the effects of cocaine-HCl were stronger and persisted longer than those of cocaine-MI. The proportion of neurons responsive to cocaine-HCl was twice that of cocaine-MI (74% and 35%, respectively). Both analogs also differed in their response onsets. Cocaine-MI rarely evoked responses after 1 min, whereas cocaine-HCl continued to evoke responses within 3 min post-injection. VTA neurons were either excited or inhibited by both cocaine analogs. Most units responsive to cocaine-MI, regardless of whether they were excited or inhibited, had electrophysiological characteristics of putative dopamine (DA) neurons. Units inhibited by cocaine-HCl also had characteristics of DA neurons, whereas excited neurons had widely varying action potential durations and discharge rates. Cocaine-MI and cocaine-HCl each produced changes in VTA neuron activity under full DA receptor blockade. However, the duration of inhibition was shortened and the number of excitations increased, and they occurred with an earlier onset during DA receptor blockade. These findings indicate that cocaine acts peripherally with a short latency and alters the activity of VTA neurons before its well-known direct actions in the brain., (Published by Elsevier Ltd.)

Mechanisms underlying cold hypersensitivity in neuropathic states are unclear. Recent data indicate both transient receptor potential (TRP) M8 and TRPA1 play a role. In relation to TRPA1, there are reported increases in mRNA. However, it is unknown whether TRPA1 mRNA is translated into functional receptors, whether these receptors are found on peripheral nociceptors and what population of primary afferents expresses the receptors. The present study provides several lines of evidence that TRPA1 receptors are expressed on intact primary sensory neurons and contribute to cold hypersensitivity following spinal nerve ligation (SNL). Immunohistochemical studies show that expression of TRPA1 is significantly increased in the ipsilateral compared with the contralateral L4 dorsal root ganglion (DRG). Using mustard oil (MO, selective TRPA1 agonist), Ca(2+) imaging demonstrates an increase in the percentage of MO-sensitive L4 DRG cells in SNL compared with sham and naive rats. The magnitude of the Ca(2+) response evoked by MO is also significantly larger in SNL compared with sham and naive rats. Behavioral studies demonstrate that SNL results in increased nocifensive behaviors to mechanical and cold stimulation that is not seen in sham or naive rats. Behavioral responses in sham rats are no different from naive rats. In vitro single fiber recordings demonstrate Adelta-fibers (intact L4 axons) in the nerve-injured hind paw have conduction velocities no different from naive rats. In contrast, compared with naive rats, mechanical thresholds of the Adelta-fibers in SNL rats are significantly decreased, the proportion of cold-sensitive and MO-sensitive Adelta-fibers is significantly increased and the response magnitude of Adelta-fibers to MO is significantly increased. MO-induced activity in Adelta-fibers is significantly reduced by Ruthenium Red (TRPA1 receptor antagonist). These results demonstrate that TRPA1 is expressed on peripheral nociceptors, and they are up-regulated on intact Adelta-fibers following nerve injury, contributing to cold hypersensitivity.

Protein kinase B (PKB/Akt) is a member of the second-messenger regulated subfamily of protein kinases. It is implicated in signaling downstream of growth factors, insulin receptor tyrosine kinases and phosphoinositide 3-kinase (PI3K). Current studies indicate that nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and PI3K help mediate inflammatory hyperalgesia. However, little is known about the role of PKB/Akt in the nociceptive system. In this study, we investigated whether PKB/Akt in primary sensory neurons is activated after noxious stimulation and contributes to pain behavior induced in rats by capsaicin. We demonstrated that phospho-PKB/Akt (p-PKB/Akt) is increased in dorsal root ganglia (DRG) at 5 min after intradermal injection of capsaicin. p-PKB/Akt is distributed predominantly in small- and medium-sized DRG cells. After capsaicin injection, p-PKB/Akt (473) is colocalized with isotectin-B4 (IB4), tyrosine kinase A (TrkA), and calcitonin gene-related peptide (CGRP). Furthermore, most transient receptor potential vanilloid type 1 (TRPV1) positive DRG neurons double label for p-PKB/Akt. Behavioral experiments show that intradermal injection of a PI3K (upstream of PKB/Akt) inhibitor, wortmannin, dose-dependently inhibits the changes in exploratory behavior evoked by capsaicin injection. The PKB/Akt inhibitor, Akt inhibitor IV, has the same effect. The results suggest that the PKB/Akt signaling pathway in the periphery is activated by noxious stimulation and contributes to pain behavior.

Included among the exciting findings in auditory neuroscience are (i) central plasticity after peripheral injury and (ii) regeneration of auditory nerve fibres following excitotoxic damage. The present study extends our understanding of auditory system plasticity by examining changes in peripheral and central physiology as the cochlea recovers from temporary deafferentation due to excitotoxicity. Application of kainic acid (60 mM) to the round window membrane substantially depressed responses from both auditory nerve and brain stem (inferior colliculus), without affecting distortion-product otoacoustic emissions from the inner ear. The auditory nerve input/output functions recovered over a 30-day period whereas recovery of brainstem response amplitudes occurred within five days. In contrast to amplitudes, thresholds at both peripheral and central levels recovered simultaneously, within five days after kainic acid application. The results indicate that (i) cochlear afferent neurons can recover after excitotoxic damage; (ii) response threshold itself, either central or peripheral, is not sufficient to assess the integrity of the auditory periphery; (iii) the central auditory system can recover more rapidly than the periphery; and (iv) the system can maintain its function in the normal range as peripheral function continues to improve.

We have previously shown that norepinephrine can produce hyperalgesia via an alpha 2-adrenergic receptor mechanism. The alpha 2-adrenergic receptor agonist clonidine has, however, also been shown to produce peripheral analgesia. In view of the multiple alpha 2-subtypes currently known (i.e. alpha 2A, alpha 2B and alpha 2C), we evaluate the alpha 2-receptor subtypes mediating norepinephrine-induced peripheral hyperalgesia and clonidine analgesia. Norepinephrine and the alpha 2-adrenergic agonists clonidine and UK 14,304 (1-1000 ng), when co-injected with the calcium ionophore A23187 (1000 ng) produced dose-dependent hyperalgesia in the Randall-Selitto paw withdrawal test. Norepinephrine (100 ng) hyperalgesia was dose-dependently antagonized by alpha 2-adrenergic receptor antagonists. From the estimated ID50, the rank order of potency was: SK&F 104856 (alpha 2B) approximately imiloxan (alpha 2B) > rauwolscine (alpha 2C) >> BRL 44408 (alpha 2A). Norepinephrine hyperalgesia was not significantly affected by pertussis-toxin treatment. Prostaglandin E2 (100 ng) hyperalgesia was inhibited dose-dependently, by clonidine and UK 14,304. Rauwolscine was more potent in reversing the inhibitory effect of clonidine on prostaglandin E2 than imiloxan while BRL 44408 was ineffective. The inhibitory effect of clonidine on prostaglandin E2 hyperalgesia was reversed by pertussis toxin. These data suggest that alpha 2B-subtype receptors mediate (norepinephrine hyperalgesia while the antinociceptive effect of alpha 2-agonist is mediated by the alpha 2C-subtype receptor. Differential coupling of these receptor subtypes to second messenger systems and location on different cell types in the rat paw may explain, at least in part, their differential responses to alpha 2-agonist stimulation, leading to hyperalgesia and analgesia.

N-methyl-D-aspartate receptors, found throughout the mammalian brain, are a component of the major excitatory transmitter system. Strong evidence exists that N-methyl-D-aspartate receptors, by promoting excessive entry of Ca2+ into neurons, play a role in neuronal damage that follows head injury, strokes, and epileptic seizures, and is associated with degenerative diseases such as Alzheimer's disease. Huntington's disease, Parkinson's disease, and amyotrophic lateral sclerosis. We have investigated whether N-methyl-D-aspartate receptors exist in peripheral neurons, and, if so, whether their activation may result in tissue injury. We report that N-methyl-D-aspartate receptors exist in the lung, that their activation triggers acute injury, and that, as in toxicity to central neurons, this injury is associated with stimulation of nitric oxide synthesis, and can be attenuated by inhibition of this synthesis. Finally, vasoactive intestinal peptide, which protects the lung and heart against oxidant injury and promotes neuronal survival and differentiation also prevented N-methyl-D-aspartate lung injury, apparently by inhibiting a key neurotoxic action of nitric oxide, but not its production. The findings suggest that N-methyl-D-aspartate receptors exist in the peripheral nervous system and that activation of these receptors, resulting in damage to peripheral neurons, may be a novel mechanism of lung and other organ injury.