Enhanced muscle fatigue occurs in male but not female ASIC3-/- mice

Abstract

Muscle fatigue is associated with a number of clinical diseases, including chronic pain conditions. Decreases in extracellular pH activates acid-sensing ion channel 3 (ASIC3), depolarizes muscle, protects against fatigue, and produces pain. We examined whether ASIC3-/- mice were more fatigable than ASIC3+/+ mice in a task-dependent manner. We developed two exercise protocols to measure exercise-induced muscle fatigue: (fatigue task 1, three 1-h runs; fatigue task 2, three 30-min runs). In fatigue task 1, male ASIC3+/+ mice muscle showed less fatigue than male ASIC3-/- mice and female ASIC3+/+ mice. No differences in fatigue were observed in fatigue task 2. We then tested whether the development of muscle fatigue was dependent on sex and modulated by testosterone. Female ASIC3+/+ mice that were ovariectomized and administered testosterone developed less muscle fatigue than female ASIC3+/+ mice and behaved similarly to male ASIC3+/+ mice. However, testosterone was unable to rescue the muscle fatigue responses in ovariectomized ASIC3-/- mice. Plasma levels of testosterone from male ASIC3-/- mice were significantly lower than in male ASIC3+/+ mice and were similar to female ASIC3+/+ mice. Muscle fiber types, measured by counting ATPase-stained whole muscle sections, were similar in calf muscles from male and female ASIC3+/+ mice. These data suggest that both ASIC3 and testosterone are necessary to protect against muscle fatigue in a task-dependent manner. Also, differences in expression of ASIC3 and the development of exercise-induced fatigue could explain the female predominance in clinical syndromes of pain that include muscle fatigue.

Fig. 1

%Change in grip force from baseline (set at 0%) immediately (Immed) and 1 h and 2 h after fatigue task 1, for the hindpaw (A—C) and the forepaw (D—F). A: change in hindpaw grip force in male acid-sensing ion channel (ASIC) ASIC3+/+ and ASIC3-/- mice after the fatigue task. *Significantly different between ASIC3+/+ and ASIC3-/- mice. B: change in hindpaw grip force in female ASIC3+/+ and ASIC3-/- mice after the fatigue task. C: change in hindpaw grip force in ASIC3+/+ and ASIC3-/- mice after various manipulations. Female mice were either ovariectomized (OVX) or ovariectomized and administered testosterone systemically (OVX, T). #Significantly less than female ASIC3+/+ mice. D: change in forepaw grip force in male ASIC3+/+ and ASIC3-/- mice after the fatigue task. *Significantly different between ASIC3+/+ and ASIC3-/- mice. E: change in forepaw grip force in female ASIC3+/+ and ASIC3-/- mice after the fatigue task. F: change in forepaw grip force in ASIC3+/+ and ASIC3-/- mice after various manipulations. Female mice were either ovariectomized or ovariectomized and administered testosterone systemically. #Significantly less than female ASIC3+/+ mice.

Fig. 2

%Change in grip force from baseline (set at 0%) immediately, 1 h, and 2 h after fatigue task 2 for the hindpaw (A—C) and the forepaw (D—F). A: change in hindpaw grip force in male ASIC3+/+ and ASIC3-/- mice after the fatigue task. B: change in hindpaw grip force in female ASIC3+/+ and ASIC3-/- mice after the fatigue task. C: change in hindpaw grip force in ASIC3+/+ mice after various manipulations. Female mice were either ovariectomized or ovariectomized and administered testosterone systemically. D: change in forepaw grip force in male ASIC3+/+ and ASIC3-/- mice after the fatigue task. E: change in forepaw grip force in female ASIC3+/+ and ASIC3-/- mice after the fatigue task. F: change in forepaw grip force in ASIC3+/+ mice after various manipulations. Female mice were either ovariectomized or ovariectomized and administered testosterone systemically.

Fig. 3

Fiber type staining in the gastrocnemius of male and female ASIC3+/+ mice. The images represent a portion of a cross section through the gastrocnemius muscle stained with one of two ATPase staining protocols, i.e., 4.3 or 4.6. In the 4.3 ATPase staining, (A and B), Type I fibers stain dark (arrows), Type IIc fibers stain intermediate (arrowheads), and Type IIa and Type IIb fibers stain light (*). In the 4.6 ATPase staining (C and D), Type I fibers stain dark (arrows), Type IIb and Type IIc fibers stain intermediate (arrowheads), and Type IIa fibers stain light (asterisks). Counting of a large sample set of ATPase-stained sections, revealed no significant gender-based differences in the percentages of fiber types.