Modulation of foregut synaptic activity controls resorption of molting fluid during larval molts of the mothManduca sexta

Abstract

The regulation and organization of a variety of stereotyped behaviors are essential for many aspects of animal development. One such collection is a series of ecdysial-related motor programs that lead to the shedding of an insect's old cuticle during a molt. Like all insects, the growth of larval Manduca sexta moths is limited by its inelastic cuticle. Consequently, growing larvae undergo a series of molts in which the old cuticle is shed and replaced by a new, larger cuticle. The two ecdysial-related motor patterns that have been the focus of much attention are the pre-ecdysis and ecdysis motor programs, which serve to loosen and then propel the old cuticle off the insect at the end of the molt, respectively (Copenhaver and Truman, 1982;Miles and Weeks, 1991; Ž itňan and Adams, 2000). The analysis of these two simple motor programs has served as a model system for the role of hormones in coordinating stereotyped behaviors. However, pre-ecdysis and ecdysis behaviors define the end of a series of ecdysial-related motor patterns that can stretch over several days. The first external sign that a Manduca larva has entered the molt cycle is apolysis, or the detachment of the old and new cuticles from one another as molting fluid (MF), a cocktail of digestive enzymes, is secreted by the epidermis into the exuvial space (the space between the old and new cuticles) where it degrades and weakens the old cuticle. Once degraded, the digested components of the exuvia and the MF are absorbed, leaving behind a thin, papery outer cuticle to be shed during ecdysis.The secretion of MF and its resorption many hours later are among a series of molt-related events that must be precisely coordinated during the molt cycle. While the removal of the MF prior to ecdysis is a feature common to all insect molts, there is no consensus on the mechanisms used to remove the MF from the exuvial space. It has often been reported that MF is resorbed across the newly formed cuticle and epidermis (Jungreis, 1979;Reynolds and Samuels, 1996;Wigglesworth, 1972). This conclusion is based on the studies of the larval molt of the bug Rhodnius prolixus (Wigglesworth, 1933(Wigglesworth, , 1972 and the pupal-adult molt of the silkworm Hyalophora cecropia (Lensky et al., 1970;Passonneau and Williams, 1953). These studies concluded that MF is absorbed into the hemocoel by passing through pores in the new cuticle.There is also evidence suggesting that the gut serves as a route for the removal of MF. Wachter (1930) We examined the role of the foregut in the resorption of molting fluid (MF) from the exuvial space during the last larval-larval molt of the moth Manduca sexta. In intermolt larvae, the activity of the foregut is characterized by robust peristaltic contractions. With the onset of the molt, MF is secreted into the exuvial space where it digests and weakens the old cuticle. The appearance of MF in the exuvial space is accompanied by a dramatic reduction in the amplitude of the foregut contractions. Foregut peristalsis returned about halfway through ...

The control of anterior foregut motility during a larval molt of the moth Manduca sexta involves the modulation of presynaptic activity

“…Recently, we found that the proper modulation of anterior foregut motility is essential for the successful completion of a larval-larval molt of M. sexta (Bestman and Booker, 2003). As described by Miles and Booker (Miles and Booker, 1994) the anterior foregut of M. sexta larvae is composed of a muscular section comprising the buccal cavity, the pharynx, esophagus, and the crop.…”

“…To gain access to M. sexta anterior foregut musculature and nervous system, larvae were dissected along the dorsal midline and the anterior foregut, brain and frontal ganglion were surgically isolated according to methods described (Bestman and Booker, 2003). The 'semi-intact' anterior foregut preparations prepared in this manner were transferred and pinned to a 35·mm plastic tissue culture dish lined with silicone elastomer (Sylgard; Dow Corning, Midland, MI, USA).…”

“…We estimated the density of FM1-43 labeled puncta by counting the number of fluorescent puncta per 20·m 2 . To monitor the effects of the inhibitory agent found in the hemolymph of early-molt stage larvae, the anterior foreguts of intermolt 5th instar larvae were first 'loaded' with the FM1-43 dye by incubating anterior foregut preparations in normal saline containing 10·m FM1-43 for 15·min (Bestman and Booker, 2003). The preparations were quickly rinsed in Ca 2+ -free saline, and then incubated for 20·min in Ca 2+ -free saline.…”

“…This blood-borne factor reduces the efficacy of the presynaptic endings of the motorneurons, resulting in a reduction in the amplitude of the excitatory junctional potential (EJP) recorded from the anterior foregut musculature. We also present evidence that crustacean cardioactive peptide (CCAP) targets theAnterior foregut modulation in Manduca sexta During a larval-larval molt, the presynaptic terminals on the anterior foregut musculature are targeted for modulation (Bestman and Booker, 2003). With the beginning of the molt, the decline in anterior foregut activity is accompanied by a sharp decline in the amplitude of the excitatory junctional potentials (EJPs) recorded from the esophageal constrictor muscles of the anterior foregut.…”

The role of the arthropod stomatogastric nervous system in moulting behaviour and ecdysis

“…This precedes the critical phase of air swallowing. Hence, the crop initially fills with moulting fluid then air (Miles and Booker, 1998;Bestman and Booker, 2003). Fluid swallowing of a different nature may also be assumed to take part in the moulting of some aquatic insects (in this case substituting for air-swallowing behaviour).…”