Activation Of ATP-sensitive Potassium Channels Antagonize Nociceptive Behavior And Hyperexcitability Of DRG Neurons

Ion channels play in vital role in the pain signal initiation and conduction. Activation of TRPV1 following a heat stimulus (>43℃) leading to pain sensation. The TRP channel family and voltage gated sodium channels are among the most intensively studied ion channels in pain signaling. Until recently, less attention has been paid to the role of potassium (K⁺) channels in pain . K⁺ channels play an essential role in setting the resting membrane potential and in controlling the excitability of neurons. Thus, K⁺ channels represent potentially attractive peripheral targets for the treatment of pain. One K⁺ channel that is known to regulate excitability in a variety of central and peripheral neurons is the M channel . Activation of the M channel by retigabine inhibits responses to the intrapaw application of carrageenan and bradykinin in rat nociceptive behavior studies. The M channel blocker XE991 evokes spontaneous pain in rats .Recently, more attention has been paid to the role of K_ATP channels in pain.The existence of K_ATP in peripheral sensory neurons and central neurons has been confirmed. In the central neurons, they regulate membrane excitability, neurotransmitter release and provide neuroprotection. It has been suggested that K_ATP may mediate the analgesic effects of morphine , clonidine and 5-HT1 agonists because the antinociceptive effects of these agents could be reversed by pretreatment with selective K_ATP antagonists but not other potassium channels blockers . Studies indicate that the nitric oxide (NO) pathway mediates the morphine activation of K_ATP .The existence of K_ATP in peripheral sensory neurons has only recently been confirmed. However, in vitro studies on DRG neurons demonstrate whole-cell K_ATP currents that either show unusual rectification properties or are expressed in only a subpopulation of the neurons . It has been suggested that K_ATP may play a major role in large diameter DRG neuron-mediated neuropathic pain. There is also evidence suggesting that the anti-nociceptive effects of K_ATP activators may stimulate mechanisms that produce anti-nociception through opioid receptor activation. However, systematic study on effects of K_ATP openers on nociception and neuronal excitability is scarce and the primary purpose of the present study is to establish the effects of direct activation of K_ATP on pain sensation and the excitability of sensory neurons.Objective: To test the effects of direct activation of K_ATP on different pain models induced by bradykinin,thermal and mechanical stimuli. For test the effects of direct activation of K_ATP on peripheral antinociception; In isoloated cultured DRG neurons, to study effects of direct activation of K_ATP on the excitability of sensory neurons.Methods: (1) Behavioral studies: Three pain models induced by bradykinin , thermal and mechanical stimuli were studied.(1) Bradykinin-induced acute spontaneous pain. The right hind paw of the animal received an intraplantar injection BK and/or K_ATP modulators, and the nocifensive responses (licking, biting, lifting and flinching) were recorded using a video camera for 30 min. The videos were analyzed by an observer unaware of the treatment allocations. (2) Mechanical withdrawal thresholds were measured using calibrated von Frey filaments applied to the plantar surface of the paw. (3) To test for thermal hyperalgesia, radiant heat was applied to the plantar surface of a hind paw from underneath a glass floor using a ray of light from a high-intensity lamp bulb. The paw withdrawal latency was recorded automatically when the paw was withdrawn from the light. 2, Electrophysiological studies: All recordings were performed using the amphotericin B perforated patch technique.Results:1 K_ATP channel openers antagonize bradykinin-induced spontaneous pain behavior through activate K_ATP channels(BK, 121±5.7 s; Pin+BK,49±5.0s; Dia+BK,59±8.0s);The antinociceptive effects of the K_ATP activators could be relieved by K_ATP blocker glyburide (Gly+Dia+BK, 104±12.4 s; Gly+Pin+BK, 85±10.8 s) and Glyburide injection alone did not induced spontaneous pain behavior(Gly, 11±8.1 s) .2 K_ATP channel openers antagonize thermal pain behavior(solvent,23±2.8 s; Pin, 36±2.9 s; Dia 35±3.0 s); Pre-injection of glyburide together with the K_ATP activators reversed the anti-thermal nociceptive effects of the K_ATP channel activators (Gly+Pin, 26±1.7 s; Gly+Dia, 31±4.1 s) and Glyburide alone did not affect thermal nociceptive behavior (26±2.5 s).3 K_ATP channel openers antagonize nociceptive response to mechanical stimuli (solvent,23±3.8g; Pin, 41±6.5g; Dia, 44±5.8g). Pre-injection of glyburide together with the K_ATP activators reversed the effects of the K_ATP channel activators (Gly+Pin, 26±4.9 s; Dia+Gly, 23±4.2 s). Glyburide alone did not affect mechanical nociceptive behavior (25±5.9 s).4 Antinociceptive effects of the K_ATP activators were the results of direct actions on the nociceptors rather than some indirect systemic effects of the K_ATP activators(Contralateral,117±11.6s,Ipsilateral,49±10.8s ); and a control vasodilator, Phentolamine, did not affect the BK-induced behavioural responses (BK,113±14.4s,Pinacidil+BK, 48±11.3 s,Phentilamine, 118±12.1s).5 Two different models of action potentials were induced by BK. One is give bradykinin increasing the frequency of action potentials and K_ATP channel opener can completely reversed the increase in action potential and the membrane potential depolarization caused by bradykinin(control, 3.7±0.85 AP/s; BK, 8.2±1.43 AP/s;BK+Pin, 2.75±0.76 AP/s. RMP; control, -50.2±3.32 mV ;BK, -42.8±3.49 mV ;BK+Pin, -52.9±3.35 mV n = 35, p < 0.01); The other is application of BK did not increase the firing frequency of the neurons, but depolarized the membrane substantially (RMP -47.5±2.81, n = 7 for control, -3.1±2.94, n = 7 for BK, p < 0.05) and, when applied pinacidil in the presence of BK, totally reversed this BK-induced depolarization ( RMP -44.3±3.28 n = 7 for BK+Pin, p < 0.05 compared with BK). 6 The K_ATP openers can moderately activate K_ATP in DRG neurons. These effects of K_ATP openers can be reversed by the K_ATP blocker glyburide. K_ATP channel openers only activate small K_ATP current in particular subpopulation of DRG neurons.Conclusion: This systematic study clearly demonstrate opening of K_ATP could have significant modulatory effects on excitability of the sensory neurons thus on sensory behavior such as nociception. The K_ATP openers diazoxide and pinacidil as a potential analgesic effect will be a hot spots in the future.