Equivalent L-type channel (CaV1.1) function in adult female and male mouse skeletal muscle fibers

Abstract

Loss of total muscle force during aging has both atrophic and non-atrophic components. The former deficit is a direct consequence of reduced muscle mass while the latter has been attributed to a depression of excitation-contraction (EC) coupling. It is well established that age-onset reductions in sex hormone production regulate the atrophic component in both males and females. However, it is unknown whether the non-atrophic component is influenced by sex hormones. Since the non-atrophic component has been linked mechanistically to reduced expression of the skeletal muscle L-type Ca²⁺ channel (Ca_V1.1), we recorded L-type Ca²⁺ currents, gating charge movements and depolarization-induced changes in myoplasmic Ca²⁺ from flexor digitorum brevis (FDB) fibers of naïve and gonadectomized mice of both sexes. Our first set of experiments sought to identify any basal differences in EC coupling or L-type Ca²⁺ flux between the sexes; no detectable differences in any of the aforementioned parameters were observed between FDB harvested from either naïve males or females. In the latter segments of the study, ovariectomy (OVX) and orchiectomy (ORX) models were used to assess the possible influence of sex hormones on EC coupling and/or L-type Ca²⁺ flux. In these experiments, FDB fibers harvested from OVX and ORX mice both showed no differences in L-type Ca²⁺ current, gating charge movement or depolarization-induced changes in Ca²⁺ release from the sarcoplasmic reticulum. Taken together, our results indicate L-type Ca²⁺ channel function and EC coupling are: 1) equivalent between the sexes, and 2) not significantly regulated by sex hormones. Since recent NIH review guidelines mandate the consideration of sex differences as a criterion for review, our work indicates the suitability of either sex for the study of the fundamental mechanisms of EC coupling. Thus, our findings may accelerate the research process by conserving animals, labor and financial resources.

Keywords: Ca(V)1.1; Excitation-contraction coupling; L-type; Orchiectomy; Ovariectomy; Sex hormones.

Fig. 1.. Equivalent EC coupling and Ca_V1.1 channel function in female and male C57BL/6 FDB fibers.

Representative myoplasmic Ca²⁺ transients elicited by 25 ms depolarizations from −80 mV to −50 through +10 mV in 10 mV increments from FDB fibers harvested from age-matched, C57BL/6J female (A) and male (B) mice. The peak ΔF/F-V relationships for naïve female and male fibers are presented in panel (C). Ca²⁺ transients were evoked at 0.1 Hz by test potentials ranging from −70 mV through +60 mV in 10 mV increments. The smooth curves in panel (C) are plotted according to Eq. 1 with the respective fit parameters shown in Table 1. Representative whole-cell recordings of L-type currents elicited by 500 ms step depolarizations from −80 mV to −40 through +50 mV in 10 mV increments are shown for female (D) and male (E) fibers. (F) Peak I-V relationships corresponding to the current families shown in panels (D) and (E). Currents were evoked at 0.1 Hz by test potentials ranging from −40 mV through +80 mV in 10 mV increments. Representative recordings of charge movements elicited by 25 ms depolarizations from −80 mV to −60 through +20 mV in 10 mV increments are shown for female (G) and male (H) fibers. (I) Q-V relationships corresponding to the charge movements shown in panels (G) and (H). Charge movements were evoked at 0.1 Hz by test potentials ranging from −70 mV through +50 mV in 10 mV increments. The smooth curves in panels (F) and (I) are plotted according to Eq. 2 and Eq. 3, respectively, with fit parameters displayed in Table 2. The numbers of analyzed fibers are indicated in parentheses. Throughout, error bars represent ±SEM.

Fig. 2.. EC coupling and Ca_V1.1 channel function in FDB fibers are little affected by ovariectomy.

Representative myoplasmic Ca²⁺ transients from an FDB fiber harvested from an OVX mouse (A). (B) the peak ΔF/F-V relationship for OVX mice (n = 7) is shown with that shown for control female mice in Fig. 1C. Representative whole-cell recordings of L-type currents are shown for an OVX fiber (C). (D) Peak I-V relationships corresponding to the current family shown in panel (C; n = 7) with the I-V relationship for control female mice from Fig. 1F. Representative recordings of intramembrane charge movements are shown for an OVX fiber (E). (F) Q-V relationships corresponding to the charge movements shown in panel (E; n = 11) with the Q-V relationship for control female mice from Fig. 1I. See Figure 1 legend for protocol details.

Fig. 3.. EC coupling and Ca_V1.1 channel function in FDB fibers are little affected by orchiectomy.

Representative myoplasmic Ca²⁺ transients from an FDB fiber harvested from an ORX mouse (A). (B) the peak ΔF/F-V relationship for ORX mice (n = 8) is shown with that shown for control male mice in Fig. 1C. Representative whole-cell recordings of L-type currents are shown for an ORX fiber (C). (D) Peak I-V relationships corresponding to the current family shown in panel (C; n = 13) with the I-V relationship for control male mice from Fig. 1F. Representative recordings of charge movements are shown for an ORX fiber (E). (F) Q-V relationships corresponding to the charge movements shown in panel (E; n = 13) with the Q-V relationship for control male mice from Fig. 1I. See Figure 1 legend for protocol details.