REGULATION OF THE FREQUENCY-DEPENDENT FACILITATION OF L-TYPE CA2+ CURRENTS IN RAT VENTRICULAR MYOCYTES

1. An increase in the rate of stimulation induces an augmentation of L -type Ca2+ currents (I-Ca) and concomitant slowing of current decay in rat ventricular cells. This facilitation is quasi immediate (1-3 s), graded with the rate of stimulation, and occurs only from negative hol ding potentials. We investigated this effect using trains of stimulati on at 1 Hz and the whole-cell patch-clamp technique (18-22 degrees C). 2. The decay of I-Ca is normally bi-exponential and comprises fast an d slow current components (I-Ca,I-fc and I-Ca,I-sc, respectively). Fac ilitation of I-Ca was observed only when I-Ca,I-fc was predominant. 3. Facilitation developed during the run-up of I-Ca with the interconver sion of I-Ca,I-fc into I-Ca,I-fc, and vanished during the run-down of I-Ca with the loss of I-Ca,I-fc. Ni2+ (300 mu M) and nifedipine (1 mu M) suppressed facilitation owing to the preferential inhibition of I-C a,I-fc. 4. Facilitation of I-Ca was not altered (when present) or favo ured (when absent) by the cAMP-dependent phosphorylation of Ca2+ chann els promoted by isoprenaline or by intracellular application of cAMP o r of the catalytic subunit of protein kinase A (C-sub). A similar effe ct was observed when the dihydropyridine agonist Bay K 8644 was applie d. In both cases, facilitation was linked to a preferential increase o f I-Ca,I-fc. 5. Following intracellular application of inhibitors of p rotein kinase A in combination with a non-hydrolysable ATP analogue, I -Ca consisted predominantly of I-Ca,I-sc and no facilitation was obser ved. The calmodulin antagonist naphthalenesulphonamide had no effect o n facilitation. 6. When Bay K 8644 was applied in combination with iso prenaline, cAMP or C-sub, the decay of I-Ca was slowed with the predom inant development of I-Ca,I-sc, and facilitation of I-Ca was nearly ab olished. Facilitation also depended on extracellular Ca2+, and was sup pressed when Ba2+ replaced Ca2+ as the permeating ion. 7. When no EGTA was included in the patch pipette, facilitation was not further enhan ced but a use-dependent decrease of I-Ca frequently occurred. When BAP TA was used in place of EGTA, the rate of inactivation of I-Ca was red uced and facilitation was abolished. 8. In conclusion, the facilitatio n of I-Ca that reflects a voltage-driven interconversion of I-Ca,I-fc into I-Ca,I-sc is also regulated by Ca2+ and by cAMP-dependent phospho rylation. The presence of the gating pattern typified by I-Ca,I-fc is required. Ca2+ may exert its effect near the inner pore of the Ca2+ ch annel protein and control the distribution between the closed states o f the two gating pathways.