Sex differences in cardiac energetics in the rat ventricular muscle

Abstract

Cardiac sex-difference functional studies have centred on measurements of twitch force and Ca²⁺ dynamics. The energy expenditures from these two cellular processes: activation (Ca²⁺ handling) and contraction (cross-bridge cycling), have not been assessed, and compared, between sexes. Whole-heart studies measuring oxygen consumption do not directly measure the energy expenditure of these activation-contraction processes. In this study, we directly quantified these energy expenditures in terms of heat production. Left-ventricular trabeculae were dissected from rats aged 9-13 weeks. Mechano-energetics of trabeculae were characterized using our work-loop calorimeter under various conditions including varying muscle lengths, stimulus frequencies, and afterloads. Each trabecula was subjected to protocols that allowed it to contract either isometrically or shorten to perform work-loops. Force production, length change, and heat output were simultaneously measured. We extracted various metrics: twitch kinetics, shortening kinetics, mechanical work, and heat associated with cross-bridge cycling and Ca²⁺ cycling, and quantified mechanical efficiency. Results show no sex differences in any of the metrics. Peak mechanical efficiency was not affected by sex (10.25 ± 0.57% in female trabeculae; 10.93 ± 0.87% in male trabeculae). We conclude that cardiac mechanics and energetics are not affected by sex at the muscle level, within the rat age range studied.

Keywords: Cardiac efficiency; Cardiac mechanoenergetics; Female; Force-length; Heat Production; Mechanical work.

Fig. 1

Representative records of a length-change protocol imposed on an isolated trabecula. (A) Records of twitches and the rate of heat production from a representative trabecula subjected to the isometric length-change at progressively diminishing muscle lengths (“i”–“iii”). Records show data of the trabecula upon electrical stimulation until steady states of force and rate of heat. (B) Twitch profiles at steady state and at various muscle lengths superimposed. Labels “i” to “iii” correspond to those in A.

Fig. 2

Steady-state isometric twitch kinetics measured under the length-change protocol. (A) Twitch duration at 5% (t₉₅) and 50% (t₅₀) of peak stress as functions of active stress of a single trabecula. Data fitted using linear regression. (B) Average relations of twitch duration at 5% (t₉₅) and 50% (t₅₀) of peak stress as functions of active stress of 17 trabeculae from female rats and those of 15 trabeculae from male rats. (C) Peak twitch duration at 5% (t₉₅) and 50% (t₅₀) of peak stress, obtained at L_o, for the female and male groups. (D) Rate of twitch stress development (rise; + dS/dt) and relaxation (fall; − dS/dt) as functions of active stress of a single trabecula. Data fitted using linear regression. (E) Average relations of ± dS/dt of 17 trabeculae from female rats and those of 15 trabeculae from male rats. (F) Peak ± dS/dt of female and male rats. No statistical differences between sex groups.

Fig. 3

Sex-difference in mechano-energetics as functions of stimulus frequency. (A) Record of the measured rate of heat production and twitch stress from a trabecula contracting isometrically under the frequency protocol. In this example, the order was as shown: 3 Hz, 5 Hz, 6 Hz, and 4 Hz. Stimulation was halted between frequency steps. (B) Average values of active stress and diastolic stress as functions of stimulus frequency. (C) Average values of twitch duration at 5% (t₉₅) and 50% (t₅₀) of peak stress as functions of stimulus frequency. (D) Average values of heat as functions of frequency. No statistical difference between sex groups can be detected.

Fig. 4

Representative example of a work-loop protocol imposed on an isolated trabecula. (A) Upon initiation of stimulation, the trabecula was required to perform isometric contractions (Isom) until a steady state was achieved; it was then required to undergo afterloaded work-loop contractions (a). This protocol was repeated for 6 afterloads (a–f). (B) Superimposed steady-state profiles of relative muscle length versus time (up-left) and overlay of steady-state isotonic twitches of varying afterloads (down-left), data from these two plots are plotted against each other to reveal work-loops in the plot on the right. The width of each loop quantifies the extent of muscle shortening. The area within each stress-length loop represents muscle work output.

Fig. 5

Shortening kinetics of trabeculae subjected to the work-loop protocol. (A) The extent of muscle shortening (calculated from the length trace in Fig. 4B) as a function of the relative active afterload of a single trabecula. Data were fitted using a 2nd order polynomial. (B) Average relation of the extent of shortening and relative active afterload of 17 female trabeculae and those of 15 male trabeculae (C) Peak extent of shortening for the muscle groups, calculated from the y-intercepts of the relations in panel (B). (D) Velocity of shortening as a function of relative afterload (calculated from the length trace as in Fig. 4B) of a single trabecula. Data were fitted using a 3rd order polynomial. (E) Average relation of velocity of shortening and relative active afterload of 17 female trabeculae and those of 15 male trabeculae from male rats. (F) Peak velocity of shortening for the muscle groups, calculated from the y-intercepts of the relations depicted in panel (E). (G) Power of shortening as a function of relative afterload of a single trabecula and (H) from the averages of 17 trabeculae from female rats and 15 trabeculae from male rats. Curves were fitted using a 3rd-order polynomials constrained to intercept the x-axis at relative afterloads of 0 and 1. (I) Peak power of shortening for the groups, calculated from the peaks of the curves in H. No differences among groups.

Fig. 6

Steady-state stresses as functions of relative length. (A) Isometric (Isom) stress (derived from Fig. 1A), work-loop (WL) stress (derived from Fig. 4A), and passive stress (derived from Fig. 1) data from a single trabecula. Data were fitted using a 3^rd order polynomial. (B) Average relations of 17 female trabeculae were not statistically different to those of 15 male trabeculae (C) Peak values of total, active, and passive stresses were not statistically difference between sex groups.

Fig. 7

Twitch heat as a function of active stress. (A) Isometric (Isom) heat (derived from Fig. 1A) and work-loop (WL) heat (derived from Fig. 4A) data from a single trabecula. Data were fitted using linear regression. Estimations of activation heat (Q_A) and peak cross-bridge heat (Peak Q_Xb) were depicted. Cross-bridge heat (Q_Xb) was quantified by subtracting Q_A from the heat measured under work-loop contractions. Peak shortening heat (Q_s) was quantified by subtracting Q_A from the y-intercept of the work-loop heat stress relation. (B) The average relations of heat and active stress of 17 female trabeculae were not different from those of 15 male trabeculae. (C) Activation heat (Q_A), peak cross-bridge heat (Peak Q_Xb), and peak shortening heat (Q_s) were not different between both sex groups.

Fig. 8

Mechanoenergetics of steady-state work-loop contractions. (A) Mechanical work (the area of each afterloaded work-loop) as a function of relative afterload from a single trabecula. Data were fitted using 3rd-order polynomial, and the peak value of work was quantified for each trabecula. (B) Average relation of work and relative afterload of 17 female trabeculae were not different from those of 15 male trabeculae. (C) Peak work was not different between sex groups. (D) Values of change of enthalpy as a function of relative afterload from a single trabecula and (E) from the averages of 17 trabeculae from female rats and 15 trabeculae from male rats. (F) The peak of change of enthalpy as a function of afterload between groups. (G) Values mechanical efficiency as a function of relative afterload from a single trabecula and (H) from the averages of 17 trabeculae from female rats and 15 trabeculae from male rats. (I) The peak of mechanical efficiency as a function of afterload between groups. Mechanical efficiency is given by the ratio of work to change of enthalpy. Data were fitted using third-order polynomials. In all panels, there were no differences among the groups.