Ca2+ entry-independent effects of L-type Ca2+ channel modulators on Ca2+ sparks in ventricular myocytes

During the cardiac action potential, Ca2+ entry through dyhidropyridine receptor L-type Ca2+ channels (DHPRs) activates ryanodine receptors (RyRs) Ca2+-release channels, resulting in massive Ca2+ mobilization from the sarcoplasmic reticulum (SR). This global Ca2+ release arises from spatiotemporal summation of many localized elementary Ca2+-release events, Ca2+ sparks. We tested whether DHPRs modulate Ca2+ sparks in a Ca2+ entry-independent manner. Negative modulation by DHPR of RyRs via physical interactions is accepted in resting skeletal muscle but remains controversial in the heart. Ca2+ sparks were studied in cat cardiac myocytes permeabilized with saponin or internally perfused via a patch pipette. Bathing and pipette solutions contained low Ca2+ (100 nM). Under these conditions, Ca2+ sparks were detected with a stable frequency of 3-5 sparks (.) s(-1) (.) 100 mu m(-1). The DHPR blockers nifedipine, nimodipine, FS-2, and calciseptine decreased spark frequency, whereas the DHPR agonists Bay-K8644 and FPL-64176 increased it. None of these agents altered the spatiotemporal characteristics of Ca2+ sparks. The DHPR modulators were also without effect on SR Ca2+ load (caffeine-induced Ca2+ transients) or sarco(endo) plasmic reticulum Ca2+-ATPase (SERCA) activity (Ca2+ loading rates of isolated SR microsomes) and did not change cardiac RyR channel gating (planar lipid bilayer experiments). In summary, DHPR modulators affected spark frequency in the absence of DHPR-mediated Ca2+ entry. This action could not be attributed to a direct action of DHPR modulators on SERCA or RyRs. Our results suggest that the activity of RyR Ca2+-release units in ventricular myocytes is modulated by Ca2+ entry-independent conformational changes in neighboring DHPRs.