RESUMO
Like seahorses, some of the closely-related pipefish species (Family Syngnathidae) incubate their eggs within a male brood pouch. This has contributed to considerable confusion about sperm transfer mechanisms to the eggs; some authors have reported that ejaculates are released directly into water before they reach the eggs, while others have suggested that eggs are fertilised using spermatozoa deposited directly into the brood pouch via an internal sperm duct. Here we present anatomical evidence from the freshwater pipefish, Syngnathus abaster, showing not only that direct sperm deposition into the pouch is impossible, but that spermatozoa must somehow travel a significant distance (>4 mm) outside the body of the male, to reach and fertilise eggs in the pouch. We have also used several putative sperm-activating solutions to identify the type of environment most conducive to sperm activation. Spermatozoa released from the testis were active for a brief period (<5 min) in water or 150 mm saline, but showed prolonged (>25 min) motility in ovarian fluid. This suggests that spermatozoa are released into a mixture of ovarian fluid and eggs while the male and female are in close contact. Our data also suggest that the fertilisation mechanism is highly efficient (sperm : egg ratio <200 : 1) even though this pipefish species produces dimorphic spermatozoa (with long and short flagellae). The shorter (<40 microm) morphotypes were not capable of motility activation, and are therefore probably incapable of fertilisation. If so, the sperm : egg ratio reported here would represent an overestimate.
