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Repeated adrenocorticotropic hormone administration alters adrenal and thyroid hormones in free-ranging elephant seals.

ABSTRACT: Understanding the physiological response of marine mammals to anthropogenic stressors can inform marine ecosystem conservation strategies. Stress stimulates the activation of the hypothalamic-pituitary-adrenal (HPA) axis and synthesis of glucocorticoid (GC) hormones, which increase energy substrate availability while suppressing energy-intensive processes. Exposure to repeated stressors can potentially affect an animal's ability to respond to and recover from subsequent challenges. To mimic repeated activation of the HPA axis by environmental stressors (or challenges), we administered adrenocorticotropic hormone (ACTH) to free-ranging juvenile northern elephant seals (Mirounga angustirostris; n = 7) once daily for 4 days. ACTH administration induced significant elevation in circulating cortisol and aldosterone levels. The cortisol responses did not vary in magnitude between the first ACTH administration on Day 1 and the last administration on Day 4. In contrast, aldosterone levels remained elevated above baseline for at least 24 h after each ACTH injection, and responses were greater on Day 4 than Day 1. Total triiodothyronine (tT3) levels were decreased on Day 4 relative to Day 1, while reverse triiodothyronine (rT3) concentrations increased relative to baseline on Days 1 and 4 in response to ACTH, indicating a suppression of thyroid hormone production. There was no effect of ACTH on the sex steroid dehydroepiandrosterone. These data suggest that elephant seals are able to mount adrenal responses to multiple ACTH administrations. However, repeated ACTH administration resulted in facilitation of aldosterone secretion and suppression of tT3, which may impact osmoregulation and metabolism, respectively. We propose that aldosterone and tT3 are informative additional indicators of repeated stress in marine mammals.

SUBMITTER: McCormley MC

PROVIDER: S-EPMC6048993 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Plasma proteome response to repeated ACTH administration in juvenile northern elephant seals. Five ul of EDTA plasma were denatured with 200 ul of 5 percent w/v SDC and 5 mM TCEP in 50 mM AmBiC at 60C for 1 hour, followed by alkylation with 20 mM CAA for 30 minutes in the dark at room temp. Samples were diluted to 0.5 percent SDC and proteins were digested using trypsin at a 1 to 50 ug enzyme to protein ratio for 16 hours at 37C. TFA was added to a final concentration of 1.0 percent to precipitate SDC, which was removed by centrifugation and extraction of the supernatant. Digested peptides were lyophilized, resuspended in 0.1 percent TFA, and desalted using Pierce Peptide Desalting Spin Columns. Eluted peptides were lyophilized and resuspended in 0.1 percent FA in LC-MS grade water. Three injections were used for each sample. For each run, 5 uL of the sample (1 ug total) was loop injected onto a reversed-phase trap column (Acclaim PepMap 100 C18 LC column; 75 um i.d. x 2 cm, 3 um particle size, 100 A pore size, Thermo Fisher Scientific, USA) by a Dionex Ultimate 3000 autosampler. Peptides were eluted onto a reversed-phase analytical column set at 35C for HPLC (EASY-SprayTM C18 LC column; 75 um i.d. x 15 cm, 100 A, Thermo Fisher Scientific, USA). Solvent A was water and B was acetonitrile (ACN), respectively (both with 0.1 percent formic acid). During each chromatographic run, which lasted 140 min, flow rates were held at 300 nl/min. Solvent B (ACN) was used as follows: 2 percent at 5 min, 2-22 percent at 75 min, 22-38 percent at 100 min, 38-95 percent at 105 min, 95 percent returning to 2 percent at 115 min, and lastly maintained at 2 percent at 140 min. Mass spectrometry analysis was performed using Orbitrap Fusion Tribrid mass spectrometer equipped with an EASY-SprayTM ion source and operated by Xcalibur 4.0 software. MS1 spectra were resolved by the orbitrap with a resolution of 120,000, scan range of 200-1400 m/z, RF lens of 60 percent, AGC target of 1.0e6, and max injection time of 50 ms. Precursor ions were isolated by quadrupole and fragmented using HCD with a collision energy of 28 percent. MS2 product ions were resolved by the orbitrap (resolution: 30,000, AGC target: 5.0e5, first mass: 100 m/z, and max injection time: 150 ms).

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Repeated adrenocorticotropic hormone administration alters adrenal and thyroid hormones in free-ranging elephant seals.

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