Effects of chronic exposure to 12‰ saltwater on the endocrine physiology of juvenile American alligator (Alligator mississippiensis)

Abstract

American alligator () habitats are prone to saltwater intrusion following major storms, hurricanes or droughts. Anthropogenic impacts affecting hydrology of freshwater systems may exacerbate saltwater intrusion into freshwater habitats. The endocrine system of alligators is susceptible to changes in the environment but it is currently not known how the crocodilian physiological system responds to environmental stressors such as salinity. Juvenile alligators were exposed to 12‰ saltwater for 5 weeks to determine the effects of chronic exposure to saline environments. Following 5 weeks, plasma levels of hormones [e.g. progesterone, testosterone, estradiol, corticosterone, aldosterone (ALDO), angiotensin II (ANG II)] were quantified using liquid chromatography and tandem mass spectrometry. Compared with freshwater-kept subjects, saltwater-exposed alligators had significantly elevated plasma levels of corticosterone, 11-deoxycortisol, 17α-hydroxyprogesterone, testosterone, 17β-estradiol, estrone and estriol whereas pregnenolone and ANG II were significantly depressed and ALDO levels were unchanged (slightly depressed). On the one hand, saltwater exposure did not affect gene expression of renal mineralocorticoid and glucorticoid and angiotensin type 1 (AT-1) receptors or morphology of lingual glands. On the other hand, saltwater exposure significantly reduced plasma glucose concentrations whereas parameters diagnostic of perturbed liver function (aspartate aminotransferase and alanine aminotransferase) and kidney function (creatinine and creatine kinase) were significantly elevated. Except for plasma potassium levels (K), plasma ions Na and Cl were significantly elevated in saltwater alligators. Overall, this study demonstrated significant endocrine and physiological effects in juvenile alligators chronically exposed to a saline environment. Results provide novel insights into the effects of a natural environmental stressor (salinity) on the renin-angiotensin-aldosterone system and steroidogenesis of alligators.

Correlations between environmental salinity levels, blood biochemistry parameters, and steroid hormones in wild juvenile American alligators (Alligator mississippiensis)

“…Hormone extractions were performed as previously described 2 , 5 , with the following modifications. Plasma samples were thawed on ice, and 500 μl aliquots of plasma were spiked with surrogate standards d9-progesterone and d3-17β-estradiol.…”

“…Chemical analyses were as described previously 2 , 5 , with the following modifications. The LC–MS/MS system comprised an Agilent 1260 UHPLC system and a triple-quad 6420 mass detector.…”

“…American alligators ( Alligator mississippiensis ) 1 do not tolerate saline environments for prolonged periods of time without access to freshwater 2 – 5 . However, many freshwater alligator habitats are prone to frequent seawater intrusion from storm surges 6 , 7 .…”

“…In addition, alligators are known to behaviorally osmoregulate by selecting lower-salinity habitats 12 , 20 , 21 . Although few studies have investigated the physiological effects of seawater on alligators, there is emerging evidence that seawater has significant impacts on alligator endocrine physiology 2 , 3 , 5 , 22 . For instance, juvenile alligators ceased feeding, experienced weight loss and showed elevated plasma corticosterone and electrolytes (Na + , Cl − ) when exposed to salinities between 10 and 20‰ for 4 weeks 3 and 5 weeks 22 .…”

“…Further, mortality was observed in juvenile alligators exposed to 15‰ and 20‰ seawater after 3 weeks 3 while 4 mortalities occurred out of 10 animals exposed to 16‰ for 5 weeks 22 . In addition, exposure of juvenile alligators to 12‰ seawater for 5 weeks significantly elevated circulating levels of electrolytes (Na + , Cl − ), lowered angiotensin II [a renin–angiotensin–aldosterone system (RAAS) hormone], and elevated important steroid hormones involved in stress response (glucocorticoids) and reproduction and growth (progestogens, androgens, estrogens) 2 . Long-term exposure to brackish seawater levels (12‰) therefore caused important alterations in reproductive ability while depressing the endocrine system responsible for water/salt regulation.…”