Sperm storage within the female reproductive tract is widespread among animal species with internal fertilization, such as the fish family Poeciliidae, wherein the females possess a sperm storage micropocket (SSP) that can store sperm for up to a year. However, the underlying mechanism and morphogenesis of SSP remain unclear. Here, using histological analysis, we found that the SSP of the western mosquitofish (Gambusia affinis) was associated with protrusions arising from the previtellogenic oocyte (PO). Similar protrusions occur in the PO of Poecilia reticulata, P. latipinna, Xiphophorus helleri, and X. maculatus, suggesting that such PO-derived protrusions might be a universal feature in Poeciliidae. These protrusions exhibit significant differences in structural composition compared to the cytoplasm. Furthermore, the area of these protrusions positively correlates with the total area of the oocytes. After maturing, the protrusions split from the oocytes, leaving behind sac-like structures composed of germinal epithelium, which later develop into the SSP for storing sperm. The bottom of the SSP is composed of a single layer of germinal epithelium. We describe and depict the protrusion formation process in the fish family Poeciliidae during the previtellogenic stage, elucidating the SSP morphogenesis.

Food scarcity is a major ecological challenge for cavefish; however, their coordinated physiological and molecular responses to prolonged starvation remain poorly understood. Here, we characterized the responses of Triplophysa rosa to 30, 60, and 90 days of food deprivation under controlled laboratory conditions by integrating growth measurements, histology, transcriptomics, quantitative reverse transcription PCR, and transmission electron microscopy. Starvation inhibited growth, reduced the condition factor and hepatosomatic index, and induced hepatocyte shrinkage, consistent with mobilization of hepatic energy reserves. Liver genes with declining temporal expression were enriched mainly in lipid biosynthesis, cholesterol metabolism, ribosomal function, and cell-cycle pathways, consistent with reduced biosynthetic activity. In the spleen, immune-related transcription increased during early starvation and declined at later stages, while the greater prominence of melano-macrophage centers was consistent with greater involvement of cellular clearance and tissue maintenance during prolonged starvation. Prolonged starvation also increased the expression of autophagy-related genes in the liver and spleen, and autophagy-related structures became more prominent in representative transmission electron microscopy images, consistent with increased involvement of intracellular recycling. Together, these findings reveal a coordinated temporal response involving metabolic adjustment, stage-dependent reorganization of splenic immune-related processes, and autophagy-associated cellular maintenance in T. rosa. This integrated approach improves our understanding of physiological maintenance during sustained nutrient limitation in cave-restricted fish.