Abstract 164: Microdomain Specific Effects of Transient Receptor Potential Channels on Pathological Cardiac Hypertrophy and Myocyte Contractility

Hypothesis: Ca2+ influx through transient receptor potential canonical (TRPC) channels and L-type Ca2+ channels (LTCCs) within caveolin-3 (Cav3) stabilized signaling microdomains provide a unique source of Ca2+ to activate pathologic cardiac hypertrophy through calcineurin (Cn)-mediated nuclear factor of activated T-cells (NFAT) signaling. We suggest that a distinct and separate population of TRPC channels localized in excitation-contraction (EC) coupling microdomains may have potent effects on myocyte contractility independent of Cav3 signaling domains. Methods and Results: Membrane localization studies and immunohistochemistry show that TRPC channels and LTCCs co-localize to Cav3 signaling microdomains. To explore a role for these caveolae based Ca2+ channels in the initiation of Cn-NFAT signaling we used an adenoviral NFAT-GFP reporter in cultured adult feline myocytes (AFMs). Infecting AFMs with ad-TRPC3 dramatically increased NFAT translocation, which was inhibited with dominant negative ad-dnTRPC6. Expression of a Cav3 targeted LTCC blocker (ad-Cav-Rem) reduced NFAT translocation while a targeted LTCC activator (ad-Cav-β2a) significantly increased NFAT activation. Neither LTCC modulator had significant effects on Ca2+ current or contractility in AFMs but we found that the expression of TRPC3 reduced myocyte contractility and induced spontaneous Ca2+ spark activity that was exacerbated by the DAG activator OAG. Moreover, dnTRPC6 blocked spontaneous Ca2+ sparks even in the presence of OAG. Immunohistochemistry analysis revealed the presence of TRPC channels in transverse tubules, consistent with the idea that they could have direct effects on EC coupling microdomains. Conclusions: Our data show that TRPC channels and LTCCs co-localize to Cav3 signaling domains where they generate a unique Ca2+ microenvironment that directly regulates Cn-NFAT signaling. Our findings also suggest that a separate and distinct population of TRPC channels within EC coupling microdomains cause reduced myocyte contractility by inducing SR Ca2+ leak and Ca2+ spark activity.