Experiment Study On The Effect Of Morphine On The L-type Calcium Channels In Neurons

ObjectiveAlough widely used in clinic,the pharmacologic mechanism, tolerance,addiction,withdrow syndrome of morphine were not clear,therefore the application were restricted.The objective of this research were to study the effect of the various concentration of morphine on the L-type calcium channels in different kinds of neurons with or without naloxone in single channel level with cell attch patch clamp tequnice,and to illustrate the mechanism of pharmacology and withdrew syndrome,and to provide the theoretical foundation for the further application in drug addiction treatment.Methods and results1.Morphine impact on the L-type calcium channels in SK-N-SH human neuroblastoma cellsMethods The SK-N-SH human neuroblastoma cells were cultured in vitro.The calcium channels in the SK-N-SH cells were recorded with cell attached patch clamp technique to analysis the physiological characteristics of the channels.Morphine were added into the bath solution with various concentration( 10-8mol/L, 10-7mol/L, 10-6rnol/L, 10-5mol/L), and the current on the calcium channels were recorded respectively.Then the cells were perincubated with naloxone(10-5mol/L),and the current on the calcium channesl were recorded with the various concentration of morphine mentioned above.Results The mean unitary slop conductance for calcium channels in SK-N-SH cells was (27.8+2.56)pS,averaged open times constants was (1.73 + 0.24)ms,ensemble averaged current was(0.107 + 0.018)pA.The open times of the channel were prolonged remarkably(P<0.01)and the ensemble averaged current were increased significantly(P<0.01),when the 0.5umol/L Bay K8644 were added into the pipette solution.These properties were in accord with the L-type calcium channels.The L-type calcium channelscurrent could be inhibited by the morphine when the concentration were up to 10-7mol/L and with the concentration dependence, nevertheless, the channel conductance were not impacted on.The inhibition could be prevented with preincubation of naloxone.2.Morphine impact on the L-type calcium channels in hippocampal pyramidal neuronsMethods Adult rat hippocampal pyramidal neurons were dissociated acutely.The effect of the various concentration of morphine on the L-type calcium channels in hippocampal pyramidal neurons were studied with cell attched patch clamp technique.Result The adult rat hippocampal pyramidal neurons acutely isolated with perfect morphology could be recorded with patch clamp technique. The L-type calcium channels in hippocampal pyramidal neurons could be inhibited by morphine when the concentration up to 10~6mol/L with the dose dependence. The effect could be blocked by naloxoen.Conclusionsl.The L-type calcium channels does exit on the human neuroblastoma SK-N-SH cell.The properties of the channel such as conductance, kinetics are similar to the other neurons.2. The current activity of L-type calcium channels could be inhibited by morphine with concentration dependence and naloxone preincubation could prevente the inhibition, hints that morphine block u-opioid receptor to inhibit the L-type calcium channels current.3.Morphine inhibited L-type calcium channels current of pyramidal neurons in rat hippocampus with concentration dependence through u-opioid receptor blocking.