Abstract
1. The gating and conduction properties of single calcium-release channels of sheep isolated cardiac junctional sarcoplasmic reticulum membranes incorporated into planar phospholipid bilayers were investigated under voltage clamp conditions at temperatures between 4 and 32 degrees C. 2. Single channel conductance was reduced linearly when temperature was decreased from 32 to 5 degrees C with a Q10 value of 1.5 between 10 and 20 degrees C. The apparent activation enthalpy for conductance between 32 and 5 degrees C was 6.16 +/- 1.2 kcal/mol. 3. Cooling the channel increased open probability (Po) when activating cytosolic calcium concentrations were varied within the range 0.1-100 microM. At an activating free calcium concentration of 10 microM, channel Po increased from 0.13 +/- 0.05 at 23 degrees C to 0.69 +/- 0.07 at 5-10 degrees C. 4. At sub-activating calcium concentrations (100 pM) or high concentrations of calcium (greater than or equal to 1000 microM), the calcium-release channel remained closed at 23 degrees C. Cooling the channel under these conditions did not increase Po. 5. Lifetime analysis indicates that with calcium as the sole activating ligand, the cooling-induced increase in Po results from an increase in channel open lifetimes with no significant alteration in the frequency of channel opening. At 23 degrees C, the open and closed lifetime distributions of the calcium-activated channel are best described by two and three exponentials respectively. At reduced temperatures (5-10 degrees C), both open and closed lifetime distributions were best described by three exponentials. 6. At sub-activating calcium concentrations, calcium-independent channel openings could be induced by sulmazole (AR-L 115 BS, 0.5-10 mM). At 23 degrees C, with sulmazole as the sole activating ligand, the best fits to both open and closed lifetime distributions were obtained with three exponentials. At reduced temperatures (5-10 degrees C), Po was increased. Gating was characterized by long open events, however the open and closed lifetime distributions were still best described by three exponentials. 7. The net effect of temperature reduction is an increase in calcium current through the channel. This finding is consistent with the suggestion that calcium release from the SR is a major factor in the initiation of rapid cooling contractures of mammalian cardiac muscle preparations.