Temperature effects on male reproductive tissues and mechanisms of recovery
Sperm presence in the SV and SV size
We scored the presence of mature sperm in both seminal vesicles (SV) and measured SV size in two- and six-day-old males as proxies for the amount of mature sperm available to males. We found a major impact of elevated developmental temperatures and the opportunity to recover on the presence of mature sperm in the SV (χ2 = 82.01,df = 4, P < 0.0001; Fig. 3A). A six-day-recovery resulted in a significant increase in mature sperm in the SV for both temperature treatments while those males not allowed to recover did not improve even after six days.
The results we observed for sperm presence were paralleled by our measure of SV size (Fig. 3B, table 2). Overall, heat-challenged males started with on average 31% smaller SVs than control males, however, by day six differences in SV size were more noticeable. Males raised at 29°C recovered to have only 15% smaller SVs compared to control males, while the other treatments retained small SV sizes resulting in a reduction of 52% for males developed at 31°C independently of ability to recover and of 43% for males raised and kept at 29°C.
We also compared sperm viability in six-day old males from the control and the 29°C recovery treatment at three timepoints after releasing sperm from SVs (Fig. 3C). This procedure allows us to determine not only sperm quality but also future sperm performance (Eckel et al., 2017). We found that, although the number of dead and alive sperm was similar at the beginning of the experiment (t0), sperm viability decreased faster over time (t15 and t30 min) when males were prior exposed to mid-challenging temperatures (GLMM with a binomial error distribution: temperature: χ2 = 16.33, df = 1,P < 0.0001; time point: χ2 = 24.22, df = 2, P < 0.0001; temperature * time point: χ2 = 9.71, df = 2, P = 0.008).