Three main topics were investigated in regulation of the titre of juvenile hormone in haemolymph of the Colorado potato beetle ( Leptinotarsa decemlineata
Say): enzymic breakdown of the hormone; binding and protection of the hormone by carrier proteins; the synthetic capacity of the corpora allata.
Juvenile hormone was broken down by two major pathways: ester hydrolysis by esterases and hydration of the epoxide group by epoxide hydratases in tissue. In haemolymph of the beetle, juvenile hormone is solely broken down by juvenile hormone esterases. An in vitro
method was developed to measure the catalytic activity of juvenile hormone esterase from haemolymph. High activities were observed in fourth-instar larvae and in beetles just before diapause. Lower activities were found in third- instar larvae and in beetles reared with long days, at diapause and after diapause. The juvenile hormone esterase was insensitive to diisopropylfluorophosphate (DFP), an inhibitor used to distinguish between carboxylesterases and esterases specific to juvenile hormone. Electrophoresis of the esterase from haemolymph showed one or more esterases specific to juvenile hormone.
The short half-life of juvenile hormone measured in vivo
and in vitro
in the haemolymph and inhibition studies with Triton X-100 suggests that juvenile hormone esterases in haemolymph govern breakdown. Activities of juvenile hormone esterase correlate well with the juvenile hormone titre.
The sharp changes in juvenile hormone esterase suggest that esterase activity is regulated. The mechanism was studied by supplying juvenile hormone and by microsurgery. Treatment of diapausing beetles with juvenile hormone itself or analogues caused an increase in activity of juvenile hormone esterase within 24 h. Ligation or removal of corpora allata suggested that this induction was an indirect effect of juvenile hormone. Transfer from short day to long day and treatment with hormone of beetles reared with short days prevented high activity of juvenile hormone esterase. Removal of corpora allata at emergence from beetles reared with short days resulted in the same. In beetles reared with short days the titre of hormone during the first days after adult emergence probably induces the rise in esterase. Esterase activity is thus most likely controlled indirectly by the hormone, via a centre in the brains (hormostate). The level of esterase activity is probably dependent on the sensitivity of this hormostate and on the titre of the juvenile hormone.
In several insects juvenile hormone is transported bound to carrier proteins. In haemolymph of larval and adult Colorado potato beetles lipoproteins of high molecular weight (>100,00 daltons) were found, capable of binding juvenile hormone, its analogues, and palmitic acid. The lipoproteins were partially separated by gel permeation chromatography and electrophoresis on polyacrylamide gel. The binding characteristics of the lipoproteins indicate low affinity (K d
M), low specificity and high binding capacity. The juvenile hormone complexed to lipoproteins was protected against esterases from haemolymph to some extent. Thus these carrier lipoproteins probably play little role in the regulation of the titre of juvenile hormone.
In the last part of our investigations the activity of the corpora allata was measured in vitro. High activities were observed in beetles reared with long days and in beetles after emergence. In beetles reared with short days, amounts of hormone produced were intermediate until Day 6 after emergence, thereafter declining to a low value. During diapause, production remained low. The production by corpus allatum and the activity of juvenile hormone esterase were in good agreement with the titre of juvenile hormone. The corpora allata are probably the primary regulator of the hormone titre in the Colorado potato beetle.