Drosophila seminal protein ovulin mediates ovulation through female octopamine neuronal signaling

Abstract

Across animal taxa, seminal proteins are important regulators of female reproductive physiology and behavior. However, little is understood about the physiological or molecular mechanisms by which seminal proteins effect these changes. To investigate this topic, we studied the increase in Drosophila melanogaster ovulation behavior induced by mating. Ovulation requires octopamine (OA) signaling from the central nervous system to coordinate an egg's release from the ovary and its passage into the oviduct. The seminal protein ovulin increases ovulation rates after mating. We tested whether ovulin acts through OA to increase ovulation behavior. Increasing OA neuronal excitability compensated for a lack of ovulin received during mating. Moreover, we identified a mating-dependent relaxation of oviduct musculature, for which ovulin is a necessary and sufficient male contribution. We report further that oviduct muscle relaxation can be induced by activating OA neurons, requires normal metabolic production of OA, and reflects ovulin's increasing of OA neuronal signaling. Finally, we showed that as a result of ovulin exposure, there is subsequent growth of OA synaptic sites at the oviduct, demonstrating that seminal proteins can contribute to synaptic plasticity. Together, these results demonstrate that ovulin increases ovulation through OA neuronal signaling and, by extension, that seminal proteins can alter reproductive physiology by modulating known female pathways regulating reproduction. In Drosophila melanogaster, the suite of seminal proteins has been identified, as have many seminal protein-dependent postmating responses, including changes in egg production and laying, remating behavior, locomotion, feeding, and in ovulation rate (reviewed in refs. 2 and 3). For example, the Drosophila seminal protein ovulin elevates ovulation rate to maximal levels during the 24 h following mating (4, 5), and the seminal protein sex peptide (SP) suppresses female mating receptivity and increases egg-laying behavior for several days after mating (6-10). However, although a receptor for SP has been identified (11), along with elements of the neural circuit in which it is required (12-14), SP's mechanism of action has not yet been linked to regulatory networks known to control postmating behaviors. Thus, a crucial question remains: how do male-derived seminal proteins interact with regulatory networks in females to trigger postmating responses?We addressed this question by examining the stimulation of Drosophila ovulation by the seminal protein ovulin. In insects, ovulation, defined here as the release of an egg from the ovary to the uterus, is among the best understood reproductive processes in terms of its physiology and neurogenetics (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27). In D. melanogaster, ovulation requires input from neurons in the abdominal ganglia that release the catecholaminergic neuromodulators octopamine (OA) and tyramine (17,18,28). Drosophila ovulation also requires an OA receptor, OA receptor in mushro...

Steroid signaling in mature follicles is important for Drosophila ovulation

“…OA binds to its receptor Octopamine receptor in mushroom body (Oamb) in mature follicle cells to induce an intracellular calcium rise, Mmp2 activation, and follicle rupture (12). Drosophila ovulation is also regulated by multiple ovarian extrinsic factors, including secretions from the oviduct (13), female reproductive glands (14), and male accessory glands (15,16). However, a role for steroid signaling in Drosophila ovulation has never been established.…”

Reproductive hacking

“…35 Since artificially increasing OA neuronal signaling compensated for lack of ovulin receipt during mating, this result suggested that ovulin increases ovulation rates by stimulating the OA pathway.…”

“…37 Consistent with this view of OA action, measures of oviduct sarcomere length showed that oviduct muscles do relax after mating. This postmating muscle relaxation depends on the female's OA signaling 35 : only females that could produce OA responded to mating by relaxing their oviduct muscles. Further, artificially and selectively increasing the activity of OA neurons using the thermally activated TrpA channel 38,39 was also able to induce the relaxation in oviduct muscles.…”

“…Further, artificially and selectively increasing the activity of OA neurons using the thermally activated TrpA channel 38,39 was also able to induce the relaxation in oviduct muscles. 35 The findings that ovulin acted through OA to increase ovulation rate and that OA affects muscle contraction in the reproductive tract suggested that ovulin and OA might together act to increase ovulation rates by regulating muscle contraction in the female reproductive tract, such as relaxing oviduct muscles or stimulating ovarian muscle contraction (although the latter is speculative). Consistent with this, the post-mating relaxation of oviduct muscles noted above did not occur if females did not receive ovulin from their mates.…”

“…Moreover, ectopically expressing ovulin in unmated females also relaxed oviduct muscles-in an OA-dependent manner. 35 There were several ways by which ovulin could regulate OA signaling. Ovulin could act upstream of OA release to increase the amount of OA neuronal activity or the number of OA synaptic release sites.…”