The immunosuppressive effects of androgens are a key component of the immunocompetence handicap hypothesis (ICHH). Here, we use bluegill sunfish (Lepomis macrochirus) to test two predictions arising from this hypothesis: (1) natural circulating concentrations of the androgen 11-ketotestosterone (11-KT) will be negatively related with measures of immunity, and (2) immune stimulation will lower circulating 11-KT concentration. We found no evidence for a relationship between natural circulating 11-KT concentration and measures of immunity (lymphocyte and granulocyte counts, respiratory burst, cytokine mRNA levels), and an immune stimulation with Vibrio vaccine did not affect circulating 11-KT concentration. We also performed a meta-analysis of immune stimulation studies to help interpret our results, and report evidence suggesting that immune stimulation has weaker effects on androgen levels in fishes compared to other vertebrates. These results suggest that the ICHH may not apply to all vertebrates, although it remains premature to state what factors account for the weaker evidence in fishes that androgens are immunosuppressive.
Resting immunity study
Field collections took place at Queen’s University Biological Station, located on Lake Opinicon (44°34′N, 76°19′W). Our first objective was to measure circulating 11-KT concentrations as well as levels of resting immunity in a wild population of bluegill (Lepomis macrochirus) during the breeding season. From June 4 to 19 2015, parental male bluegill (n = 24) were collected either by angling in pelagic areas or with dip nets in littoral areas while males were guarding nests that contained few or no eggs (≤10% of the nest surface covered by eggs, such that males were assumed to be in the early stages of spawning). For all fish, 400 μL of whole blood was drawn from the caudal vessel using a heparinized syringe. The fish were then measured for total body length and released. The time from collection to the end of blood sampling was recorded for each fish (always <2 min from capture). For the respiratory burst assay, 200 μL of whole blood was then transferred into tubes containing 20 μL of 600 units/mL ammonium heparin. The remaining 200 μL of blood was transferred into non-heparinized tubes for blood smear cell counts (lymphocyte and granulocyte) and hormone analyses. Blood samples were kept on ice after collection and then stored at −20 °C prior to 11-KT analysis.
Immune stimulation study
To test whether circulating 11-KT concentrations decline following immune stimulation, on June 15, 2015 we captured a group of parental males (n = 11) using dip nets while they were guarding nests that contained few or no eggs. We measured total body length and clipped a unique combination of dorsal spines on each individual to allow subsequent identification. We then transferred each individual to a separate 60 × 60 × 60 cm floating net pen situated in the littoral zone of the lake. Previous trials have shown that floating net pens support confinement of bluegill for up to 14 days with low mortality and no elevation of glucose levels that would indicate an energetic response to a stressor (Gutowsky et al., 2015).