Shedding light on post-copulatory sexual selection

Fertilisation requires one sperm. The selection of the 'winning' sperm out of the millions inseminated into the female is not determined by a simple lottery, but rather a complex set of interactions between sperm and the female reproductive tract. These post-mating interactions, collectively termed post-copulatory sexual selection (PCSS), are influenced by many factors from biochemical to behavioural, making PCSS dynamic, complicated, and challenging to study. In some taxa, including birds, sperm take a surprising diversion in their long journey between copulation and conception. Females of many species can store sperm for extended periods of time in specialised storage sites within their reproductive tracts. Such storage can give females the opportunity to bias fertility outcomes through PCSS mechanisms. However, these processes occur deep within the female body, making them particularly difficult to document. A critical step towards understanding these mechanisms involves visualising them in real time, inside female tissue, under physiological conditions that are as close to natural as possible. Such visualisation is not trivial - especially in birds - due to the challenges of imaging through thick, muscular tissue. In this thesis, I demonstrate the optimisation of selective plane illumination microscopy (SPIM) and the development of imaging protocols, including sample preparation and cell labelling for this novel application. SPIM is a light sheet fluorescence microscopy technique, which allowed me to optical section live oviduct tissue to reveal the three-dimensional structure of sperm storage tubules (SSTs) in zebra finch [Taeniopygia guttata] females. This imaging technique, in combination with electron microscopy, also allowed me to make volumetric observations of zebra finch sperm mid-piece structure. These studies have helped us understand the adaptive significance of these structures, as well as the functioning of SSTs and the possible sperm-female interactions that occur inside them.