ryanodine and Pain

Propofol, a general anesthetic administered intravenously, may cause pain at the injection site. The pain is in part due to irritation of vascular endothelial cells. We here investigated the effects of propofol on Ca2+ transport and pain mediator release in human umbilical vein endothelial cells (EA.hy926). Propofol mobilized Ca2+ from cyclopiazonic acid (CPA)-dischargeable pool but did not cause Ca2+ release from the lysosomal Ca2+ stores. Propofol-elicited Ca2+ release was suppressed by 100 μM ryanodine, suggesting the participation of ryanodine receptor channels. Propofol did not affect ATP-triggered Ca2+ release but abolished the Ca2+ influx triggered by ATP; in addition, propofol also suppressed store-operated Ca2+ entry elicited by CPA. Ca2+ clearance during CPA-induced Ca2+ discharge was unaffected by a low Na+ (50 mM) extracellular solution, but strongly suppressed by 5 mM La3+ (an inhibitor of plasmalemmal Ca2+ pump), suggesting Ca2+ extrusion was predominantly through the plasmalemmal Ca2+ pump. Propofol mimicked the effect of La3+ in suppressing Ca2+ clearance. Propofol also stimulated release of pain mediators, namely, reactive oxygen species and bradykinin. Our data suggest propofol elicited Ca2+ release and repressed Ca2+ clearance, causing a sustained cytosolic [Ca2+]i elevation. The latter may cause reactive oxygen species and bradykinin release, resulting in pain.

Topics: Adenosine Triphosphate; Bradykinin; Calcium; Endothelial Cells; Humans; Pain; Propofol; Reactive Oxygen Species; Ryanodine; Ryanodine Receptor Calcium Release Channel

Tetanic stimulation of the sciatic nerve induces long-term potentiation (LTP) of C-fiber-evoked field potentials in the spinal dorsal horn and persistent pain, suggesting that spinal LTP may be a substrate for central sensitization of the pain pathway. However, its cellular mechanism remains unclear. The present study provides electrophysiological and behavioral evidence for the involvement of ryanodine receptor (RyR) in the induction of spinal LTP and persistent pain in rats. The specific inhibitor of ryanodine receptor, ryanodine and dantrolene, dose dependently blocked the induction, but not maintenance, of spinal LTP and reduced persistent pain behaviors induced by tetanic sciatic stimulation. Both cyclic ADP ribose (cADPR), an endogenous agonist of RyR, and (±)-1,4-dihydro-2,6-dimethyl-5-nitro-4-[2-(trifluromethyl)-phenyl]-3-pyridine carboxylic acid methyl ester (Bay K 8644), an agonist of L-type calcium channel, attenuated ryanodine-induced inhibition. Immunohistochemistry and electron microscopic observation showed that RyR subtypes RyR1 and RyR3 were located in the spinal dorsal horn. The results suggest that RyRs are involved in synaptic plasticity of the spinal pain pathway and may be a novel target for treating pain. © 2012 Wiley Periodicals, Inc.

Topics: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; Analysis of Variance; Animals; Biophysics; Calcium Channel Agonists; Dantrolene; Disease Models, Animal; Dose-Response Relationship, Drug; Electric Stimulation; Evoked Potentials; Functional Laterality; Gene Expression Regulation; Long-Term Potentiation; Male; Microscopy, Electron, Transmission; Muscle Relaxants, Central; Nerve Fibers, Unmyelinated; Pain; Pain Measurement; Posterior Horn Cells; Rats; Rats, Sprague-Dawley; Ryanodine; Ryanodine Receptor Calcium Release Channel; Sciatic Nerve; Spinal Cord

Calcium (Ca++) administered into the i.c.v. space of mice has been reported to block opioid-induced antinociception dose dependently. These studies were conducted to test the hypothesis that Ca++ i.c.v. blocks the antinociceptive effects of morphine i.c.v. as a consequence of transmembrane Ca++ influx and Ca++ release from intracellular pools. Mice were injected with voltage-sensitive Ca++ channel antagonists at a dose that did not affect morphine antinociception to determine whether this pretreatment would prevent the inhibitory effects of Ca++. Nimodipine (12 nmol i.c.v.) was ineffective in preventing the inhibitory effects of Ca++ (100 nmol i.c.v.), whereas omega-conotoxin GVIA (3.3 pmol i.c.v.) completely prevented the inhibition by Ca++ of morphine antinociception. Other experiments were conducted to determine whether blocking Ca++ release from Ca++/caffeine-sensitive microsomal pools with ryanodine would prevent the inhibitory effects of Ca++. Ryanodine (2 nmol i.c.v.) significantly attenuated the inhibition by Ca++ of morphine antinociception. Another hypothesis to be tested was that stimulation of Ca++ release from intracellular pools would, like Ca++, block morphine antinociception. Thapsigargin (0.002-30 nmol i.c.v.), which increases cytosolic Ca++ by depleting Ca++ from inositol 1,4,5-trisphosphate-sensitive microsomal pools, dose-dependently blocked the antinociceptive effects of morphine. The results of this study indicate that Ca++ blocked morphine antinociception by stimulating Ca++ influx through omega-conotoxin GVIA-sensitive channels and by stimulating Ca++ release from Ca++/caffeine-sensitive microsomal pools.

Topics: Animals; Caffeine; Calcium; Calcium Channels; Cell Compartmentation; Male; Membrane Potentials; Mice; Mice, Inbred Strains; Microsomes; Morphine; Nimodipine; omega-Conotoxin GVIA; Pain; Peptides; Ryanodine; Terpenes; Thapsigargin

Bradykinin is one of several pro-inflammatory, pain-inducing substances produced during inflammation--the body's response to injury. In previous work we have shown that bradykinin and guanosine-5'-O-3-thiotriphosphate increase excitability in a subpopulation of cultured neonatal rat dorsal root ganglion neurons. We now describe experiments in which the mechanism underlying the stimulatory action of these two substances has been examined in more detail. Using the whole-cell voltage-clamp technique, bradykinin-sensitive cells were distinguished by their response to a 1-s depolarizing voltage-pulse which evoked more than one inward current during the step command. The secondary inward currents are likely to represent action potentials generated at the poorly clamped neurites of these cells. Bradykinin- and guanosine-5'-O-3-thiotriphosphate-induced changes in excitability were measured indirectly by a change in the number of inward currents recorded during the 1-s depolarizing voltage-step. The effect of activators and inhibitors of protein kinase C, arachidonic acid metabolism, G-protein activation and release of intracellular Ca2+ were examined on this response. In the presence of extracellular staurosporine (1.0 microM) or nordihydroguaiaretic acid (10 microM), these excitatory effects were reduced but not abolished, whilst indomethacin (20 microM) had no effect. Intracellular application of guanosine-5'-O-2-thiodiphosphate (10 mM) or ryanodine (100 microM) substantially reduced the effect of bradykinin. The excitatory effect of internal guanosine-5'-O-3-thiotriphosphate (500 microM) occurred gradually over time, and this was mimicked by internal application of myo-inositol 1,4,5-trisphosphorothioate (1.0 microM). From the results, it is proposed that G-protein activation is an essential component of the bradykinin response, which may also require a Ca(2+)-activated conductance modulated by protein kinase C and lipoxygenase metabolites of arachidonic acid.

Topics: Alkaloids; Animals; Animals, Newborn; Bradykinin; Cells, Cultured; Evoked Potentials; Ganglia, Spinal; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Indomethacin; Kinetics; Masoprocol; Neurons, Afferent; Pain; Protein Kinase C; Rats; Rats, Wistar; Ryanodine; Signal Transduction; Staurosporine