Previously, we analyzed contractile function of aged murine ventricular myocytes. We showed that contraction was slowed in aged ventricular myocytes. We found NADPH oxidase subunits and NOS1 to be upregulated in aged hearts suggesting a role for superoxide and nitric oxide in cardiac aging. Accordingly, contraction of aged myocytes was accelerated by scavenging superoxide with tiron and NOS1 deficient mice showed no alterations in contractile function with aging.

Here, we analyzed calcium transients of ventricular myocytes isolated from young (2–4 months) and aged (24–26 months) mice by means of indo-1 fluorescence ratio. Isolated myocytes were continuously stimulated by voltage clamp pulses from -80 to -45 mV (prepulse, 10 ms) immediately followed by a voltage step to 0 mV (160 ms) at 1Hz (37°C).

In aged wild-type ventricular myocytes, the amplitude of calcium transients was significantly increased (young: 231 18 a.u., n = 32; aged: 307 26 a.u., n = 24; mean S.E.M.). This contrasts with an unchanged fractional shortening of aged ventricular myocytes. Furthermore, although time to peak shortening and time to 70% relengthening were significantly prolonged in aged ventricular myocytes, concerning calcium transients only time to 70% calcium decay was affected in aged myocytes (young: 196 6 ms; n = 32; aged: 237 7 ms, n = 24).

The increased calcium transient amplitude of aged myocytes was normalized by tiron (aged + tiron: 206 32 a.u., n = 17). However, prolonged time to 70% calcium decay was unaffected by superoxide scavenging (aged + tiron: 225 8 ms, n = 17). Calcium transients of ventricular myocytes derived from aged NOS1 deficient mice showed neither increased amplitudes (aged NOS1^-/-: 236 12 a.u., n = 22) nor prolonged time to 70% calcium decay (191 7 ms).

We conclude that superoxide and nitric oxide derived from age-dependently upregulated NADPH oxidase and NOS1 causally contribute to the contractile dysfunction of aged murine ventricular myocytes. These reactive oxygen and nitrogen species modulate the calcium homeostasis of aged myocytes, but also modulate the function of the myofilaments, i.e. by modulation of calcium sensitivity.