The role of low levels of juvenile hormone esterase in the metamorphosis of Manduca sexta

Abstract

The activity of juvenile hormone esterase (JHE) in feeding fifth instar larvae of Manduca sexta increases gradually with larval weight and rises to a peak after larvae pass the critical weight when juvenile hormone secretion ceases. Starvation of larvae of Manduca sexta (L.) that had exceeded the critical weight inhibited peak levels of JHE, but did not delay entry into the wandering stage when larvae leave the plant in search of a pupation site. This suggests that peak levels of JHE may not be essential for the normal timing of metamorphosis. Starved larvae pupated normally, indicating the peak of JHE was not necessary for a morphologically normal pupation. Treatments of larvae with the selective JHE inhibitor O-ethyl-S-phenyl phosphoramidothiolate (EPPAT) that began immediately after larvae achieved the critical weight (6.0 to 6.5 grams for our strain of Manduca) delayed entry into the wandering stage. By contrast, EPPAT treatment of larvae at weights above 8.0 g had no effect on the subsequent timing of the onset of wandering. Therefore, although the normal timing of the onset of wandering does not require peak levels of JHE, it requires low to moderate levels of JHE to be present until larvae reach a weight of about 8.0 g.

Figure 1.

The juvenile hormone esterase (JHE) activity profile of normally feeding last (5^th) instar larvae of Manduca sexta. JHE activity was measured as nmoles of JH-acid produced per minute per ml of hemolymph. Each point is the mean of 9 to 11 pre-wandering larvae, or 4 to 6 post-wandering larvae. Vertical bars are standard errors of the mean (which in some cases do not exceed the size of the symbol). The abscissa is a measure of the physiological age of the larva, which is given on two scales: before entry into the wandering stage the larva's physiological age is given by its weight, and after entry into the wandering stage it is given in days. The curve represents the best-fit polynomial regression (2^nd degree polynomial, r² = 0.99), and is extrapolated to animals weighing 12 grams (the maximal weight at which larvae entered the wandering stage).

Figure 2.

The effect of starvation after the critical period on the JHE activity profile of 5^th instar Manduca sexta larvae. JHE activity was measured as nmoles of JH-acid produced per minute per ml of hemolymph. Each point is the mean of 7 to 9 pre-wandering larvae, or 4 to 6 post-wandering larvae. Vertical bars are standard errors of the mean (which in some cases do not exceed the size of the symbol). Larvae were starved at weights of 7 ± 0.5 grams (dark vertical bar), and JHE activity was measured at 24-hour intervals thereafter. The expected JHE activity of normally feeding larvae (from Fig. 1) is indicated by the dotted line.

Figure 3.

Effect of EPPAT treatment on the timing of entry into the wandering stage of normally feeding Manduca sexta larvae. The abscissa indicates the weight at which EPPAT treatment began. P-values above each set indicate the probability that experimental and control values are identical (Wilcoxon's signed-ranks test; N = 10 for each bar). Vertical lines are standard deviations.