The amplitude and time course of the myoplasmic free [Ca2+] transient in fast-twitch fibers of mouse muscle

Abstract

Bundles of 10-100 fibers were dissected from the extensor digitorum longus muscle of mouse, mounted in an apparatus for optical recording, and stretched to long sarcomere length (≤ 3.6 μm). One fiber within the bundle was microinjected with furaptra, a fluorescent indicator that responds rapidly to changes in myoplasmic free [Ca2+] (Δ[Ca2+]). Twitches and brief tetani were initiated by external stimulation. At myoplasmic furaptra concentrations of ~0.1 mM, the indicator's fluorescence signal during fiber activity (ΔF/F) was well resolved. ΔF/F was converted to Δ[Ca2+] under the assumption that furaptra's myoplasmic dissociation constant for Ca2+ is 98 μM at 16°C and 109 μM at 28°C. At 16°C, the peak amplitude of Δ[Ca2+] during a twitch was 17.8 ± 0.4 μM (±SEM; n = 8) and the half- width of Δ[Ca2+] was 4.6 ± 0.3 ms. At 28°C, the peak and half-width values were 22.1 ± 1.8 μM and 2.0 ± 0.1 ms, respectively (n = 4). During a brief high-frequency tetanus, individual peaks of Δ[Ca2+] were also well resolved and reached approximately the same amplitude that resulted from a single shock; the initial decays of Δ[Ca2+] from peak slowed substantially during the tetanus. For a single twitch at 16°C, the amplitude of Δ[Ca2+] in fast-twitch fibers of mouse is not significantly different from that recently measured in fast-twitch fibers of frog (16.5 ± 0.9 μM; Zhao, M., S. Hollingworth, and S.M. Baylor, 1996. Biophys. J. 70:896-916); in contrast, the half-width of Δ[Ca2+] is surprisingly brief in mouse fibers, only about half that measured in frog (9.6 ± 0.6 ms). The estimated peak rate at which Ca2+ is released from the sarcoplasmic reticulum in response to an action potential is also similar in mouse and frog, 140-150 μM/ms (16°C).