Effect of temperature on the activation of myocardial KATP channel in guinea pig ventricular myocytes:: a pilot study by whole cell patch clamp recording

Background The myocardial ATP sensitive potassium channel (K-ATP channel) has been known for more than two decades, the properties of this channel have been intensively investigated, especially the myocardial protection effect by opening this channel. Numerous studies, including hypothermic, using K-ATP agonists to achieve a hyperpolarizing cardioplegic arrest, have shown a better myocardial protection than potassium arrest. However, there is no evidence showing that K-ATP channel could be opened by its agonists under profound hypothermia. We investigated the effect of temperature on activation of myocardial K-ATP channel by nicorandil. Methods Isolated ventricular myocytes were obtained by collagenase digestion of the hearts of guinea pigs and stored in KB solution at 4 degrees C. With a steady ground current, the myocytes were perfused with 1 mmol/L nicorandil until a steady IKATP occurred. Then the cells were perfused with 1 mmol/L nicorandil plus 1 mu mol/L glybenclamide. Currents signals were recorded on whole cells using patch clamp technique at several temperatures. The temperature of the bath solution around myocytes was monitored and was controlled at 4 degrees C, 10 degrees C, 20 degrees C, 25 degrees C and 35 degrees C respectively. About 10 cells were tested at each temperature, the cells were considered useful only when the outward current could be induced by nicorandil and blocked by glybenclamide. All data were analyzed using Graphpad PRISM 3.0 (Graphpad, San Diego, CA, USA). Nonlinear curve fitting was done in Clampfit (Axon) or Sigmaplot (SPSS). Results At 4 degrees C, 10 degrees C, 20 degrees C, 25 degrees C and 35 degrees C, the time needed to open the myocardial K-ATP channel was (81.0 +/- 0) minutes, (50.5 +/- 11.7) minutes,. (28.8 +/- 2.3) minutes, (9.4 +/- 10.2) minutes and (2.3 +/- 1.0)minutes respectively (P=0.003). The linear relationship between temperature and time needed to open the channel was y (min) (4348.790-124.277x)/60, where y (min) is time needed to open K-ATP channel, x is temperature, correlation coefficient r =-0.942 (P=0.00), regression coefficient b =-124.277 (P=0.00). The current densities among different temperatures were statistically different (P=0.022), the current density was greater after the activation of K-ATP channel at higher temperatures. The lower the temperature, the fewer cells in which K-ATP channels could be opened. At 4 degrees C, only one cell in which the K-ATP channel could be opened, took a quite long time (81 minutes) and the I-V curve was quite untypical. Conclusions K-ATP channel activated by temperature linearly related to time needed to open K-ATP channel; the lower the temperature, the longer the time needed to open channel and the smaller the current density. At profound hypothennia, it is difficult to activate K-ATP channels.