Do Metabolic Enzymes Explain Osmoregulation in Juvenile American Eels

During the spring, juvenile eels ( Anguilla rostrata ) (5‐6 cm; 50‐200 mg) gather in creeks along the Hudson River Tributary Estuary after migration from the Atlantic Ocean. This requires them to quickly adapt to environments with lower salt concentrations. We wondered how this is achieved. To study eels in a lab setting, we transferred them into Instant Ocean® sea salt (100% artificial sea water [ASW]) or dilutions thereof for 2 weeks; mortality was only observed in 100% ASW (37% expired in 3 hours). Interestingly, after 4 hours, 10 or 50% ASW eels showed little change in body mass, suggesting osmoregulation. While much progress has been made in understanding the mature eel response, few studies have reported enzyme activities in young eels. We hypothesized that dehydrogenases and acetylcholine esterase activities might provide a clue to explain osmoregulation and survival. Supernatants of whole juvenile eel extracts obtained from these experiments (IACUC‐approved protocol) were run on 7.5% Tris‐glycine PAGE gel (150V, 50 min) and the activities of non–denatured isozyme bands were analyzed with Bio‐Rad ChemiDoc™ XRS+ Imager. Denatured supernatant extracts were also run on TGX 4‐15% Tris‐glycine PAGE gel to assess total protein banding. There was little difference in number of protein bands between eels exposed to 10% vs 50% ASW(2 and 3 weeks); however, band intensities were significantly higher for 50% ASW. Further studies revealed that two MDH (malate dehydrogenase) isozyme bands were present in extracts of eels exposed to a broad range of salinities: a relatively heavier band as mitochondrial (m) MDH and a lighter cytoplasmic (c) MDH. Activities of the mMDH bands were more robust than the cMDH bands at 50% and 10% ASW. Although the mMDH activities in eels exposed to either salinity were similar, the cMDH activities in eels exposed to 50% ASW were greater than in eels exposed to 10%. Additional experiments identified four acetylcholine esterase (AChE) isozyme bands in eel extracts; the activity of only one was significantly prominent in all exposures. Transferring the eels from 50% to 10% ASW reduced activity of the relatively heaviest isozyme band. Notably, we could not discern any fluctuations in lactate dehydrogenase (LDH) levels. In conclusion, while these preliminary findings do not reveal any concrete trends in metabolic regulation tied to survival, it does appear several key enzymes have a translational correlation tied to saline homeostasis. We acknowledge that the interpretation of these experimental results is complicated by a lack of genetic validation (i.e., sequences information and parental lines) ‐ “nature” . In addition, the investigators were unable to track or define the environmental stresses faced during migration by individual juvenile eels before capture, which may manifest itself in unique stress responses ‐ “nurture.”