Propofol Shifts Ca²⁺ Sensitivity And Kinetics Of Bk_Ca In Mouse Cerebral Arterial Smooth Muscle Cells

Abstract: Objective: Propofol is widely used for intravenous anesthesia with the side effect of hypotension. Many mechanisms are involved in the hypotension, including: inhibition of the renin angiotensin system; decreasing sympathetic nerve activity, inhibiting the noradrenaline release from the sympathetic nerves; inhibitory effect of the vasomotor centre; decreasing the myocardial contractility and cardiac output; inducing the peripheral vascular relaxation directly and the most important mechanism is the direct effect on peripheral vascular. Large conductance Ca²⁺-sensitive K⁺ channels (BK_Ca) are broadly expressed on vascular smooth muscle cells and take 70-80% outward current of the whole cell. The activation of BK_Ca induces cell membrane hyperpolarzation which inhibits L type voltage-dependent calcium channels, decreases calcium ion concentration in cytoplasm and then causes vascular dilatation. Because of its high conductance and broadly expression on vascular smooth muscles, BK_Ca play an important role in regulating the vascular tone and they are one kind of the targets of the vasoactive agents. Previous studies suggested that the activation of large conductance BK_Ca was involved in propofol-mediated hypotension, and some researchers confirmed that propofol activated BK_Ca using patch clamp technique. However, the underlying mechanism of the activation, especially the kinetics and its target, remains unknown. Here, we aim to investigate the effect of propofol on kinetics and Ca²⁺ sensitivity of BK_Ca. Methods: Mice (15-20 g) were anesthetized by intraperitoneal injection 60 mg/kg napental and then were decapitated. Brains were taken out and put in ice pretreated physiological saline solution (PSS) and cerebral arteries were isolated from pavimentum cerebri. Smooth muscle cells were isolated from mouse cerebral arteries after two steps of enzymatic digestion and then stored at 4℃. BK_Ca macro-currents and spontaneous transient outward K⁺ currents (STOCs) were recorded under different propofol concentrations in perforated whole-cell configuration. We recorded Ca²⁺ sensitivity, voltage dependence and kinetics of BK_Ca under different propofol concentrations in inside-out single channel configuration. Results:â‘ The characteristic properties of BK_Ca in the mouse cerebral arterial smooth muscle cells, i.e. large conductance, voltage dependent- and calcium dependent- characteristics, are very similar to those found in other tissues.â‘¡Propofol increased the BK_Ca macro-currents and spontaneous transient outward K⁺ currents (STOCs). At Vm= +60 mV, 56μM and 112μM propofol increased the macro-current density 1.44-fold and 2.44-fold, respectively. At Vm= -30 mV, 56μM propofol increased the amplitude of STOCs 1.51-fold and frequency 1.69-fold.â‘¢Propofol increased the total open probability (NPo) of single BK_Ca in a concentration-dependent manner. 56μM and 112μM propofol increased NPo 52-fold and 193-fold with the half maximal effective concentration (EC50) of 75.93μM. Propofol shifted the NPo-[Ca²⁺]free curve left with the equilibrium dissociation constant (K_d) of Ca²⁺ from 0.887μM (control) to 0.694 (28μM propofol), 0.599μM (56μM propofol) and 0.176μM (112μM propofol).â‘£Analysis of the channel kinetics revealed that 112μM propofol increased the open dwell time and decreased the closed dwell time significantly. It decreased the open time constant and increased the closed time constant. Conclusion: Propofol activates BK_Ca through increasing the calcium sensitivity and voltage dependence of the channels. Further more, propofol modulates the kinetics including open and closed states, and inhibits the transmit from the open state to close state and fastens the transmit from the close state to open state.