Jh 3

Females were treated with either JH III (Sigma) or methoprene (Zoecon, Palo Alto, CA). Both JH III and methoprene were dissolved in cyclohexane (HPLC grade, Sigma-Aldrich) at final concentrations of 10 μg/μl each and 1 μl was topically applied to the gaster of cold-anesthetized wasps. Control wasps received a solvent application, and in some cases, non-treated wasp controls were also included. To test for immediate behavioral responses, 10 μg of JH III was applied on three consecutive days to old (>3 weeks) queenless workers and, in another experiment, to 1–3 day old adult females who emerged as the colony was transitioning from being queenless to queenright. To assess JH effects on ovarian growth and chemical signaling, 10 μg of methoprene, which has longer-lasting effects than JH III [47 (link)], was applied to working wasps at 3 and 6 days following eclosion. All wasps were kept in isolation for 45–60 min before they were returned to the nest to reduce the spread of the topically applied hormone to nestmates. Unless otherwise noted, all nests containing treated females were observed for 1–2 hours each day before treatment or capture.

A 1 μg aliquot of JH-III (Sigma-Aldrich) dissolved in 0.5 μl of acetone was applied to the abdomen of one-day-old females. Control females were treated with acetone (0.5 μl). 10 h after application, half of the JH- and acetone-treated flies were exposed to heat stress and then all of the flies were frozen in liquid nitrogen and stored at −20°C.