Neuropeptides are responsible for controlling different aspects of physiology and behavior including body growth, eating, drinking, sex, etc. The broad goal of this proposal is to understand cellular mechanisms controlling synthesis of a neuropeptide hormone using neurons that control reproduction in the mollusk Aplysia. Earlier studies from this laboratory showed that electrical excitation of the reproductive neuroendocrine bag-cell neurons (BCNs) led to large amounts of secretion of the neuropeptide egg laying hormone (ELH). The releasable amount of ELH was replenished by upregulating ELH synthesis at the expense of most other proteins in the BCNs. It was revealed that electrical excitation of the BCNs stimulated a novel protein-synthetic pathway involving a special code on ELH mRNA called an internal ribosomal entry site (IRES). This is similar to how viruses control protein synthesis in cells following infection, and was not thought to play a physiological role in cellular systems. The aim of the proposed studies is to understand how stimulation of the BCNs activates the IRES on ELH mRNA. Based on earlier work, it is hypothesized that the cyclic AMP second messenger pathway (via cyclic AMP response-element binding protein, or CREB) is playing an important role in mediating the effects of electrical excitation on IRES activation, leading to upregulation of ELH synthesis. To test this hypothesis, expression of CREB will be obstructed by RNA inhibition, and the effects on electrical excitation of ELH synthesis and IRES activation will be documented. This experiment uses a combination of molecular, biochemical, electrophysiological, and optical imaging techniques. It is expected that inhibition of CREB will prevent electrical activation of the IRES and subsequent upregulation of ELH synthesis. This work is at the forefront of a completely new way of thinking about the way that cellular systems can regulate synthesis of proteins
