Effects of glucoprivation on the adrenal medulla: a time course study using shotgun proteomics

Abstract

Theoretical thesis.Bibliography: pages 55-65.1. Introduction -- 2. Experimental procedures -- 3. Results -- 4. Discussion.The adrenal medulla is the sole endocrine gland responsible for catecholamine release inresponse to stress. While a time course of stress response has been described for rate-limitingenzyme in catecholamine synthesis; tyrosine hydroxylase (TH), none have yet described any other metabolic changes in this gland. Therefore the aims of this study are to map the proteome of the adrenal medulla and determine any changes over time following an acute episode of glucopenia, a potent stimulus thereof. In addition, the final aim is to determine whether or not Western blot analysis reflects the findings shown using mass spectrometry (MS). A single IP injection of 2-deoxyglucose was used to induce glucopenia in Sprague Dawley rats. The adrenal glands were harvested over at 0min, 20min, 50min, 4h ,8h and 24h after the stimulus (n=6 for each timepoint). Label-free LC-MS/MS was then performed on the samples (n=3) for each timepoint to obtain the proteomic profile. Profiles were quantitatively compared to the control (0min/unstimulated). Western blotting was then used in order to validate the MS data using specifically selected proteins. The protein profile was determined and comprises 973 proteins, with marked changes at each time point; 20min=78, 50min=138, 4h=163, 8h=136 and 24h=107. Western blot analysis regarding TH corresponded with MS data, however other proteins showed differing expression levels. From this we understand that cellular metabolic changes occur rapidly in response to stress and last longer than 24h. It is also apparent that Western blot analysis does not always correspond with the MS output.Mode of access: World wide web1 online resource (iv, 106 pages) diagrams, graphs, table