EFFECT OF LONG-TERM INSULIN EXPOSURE ON INSULIN BINDING IN TETRAHYMENA-PYRIFORMIS

Previous studies have indicated that Tetrahymena pyriformis can bind t he vertebrate hormone insulin. Conventional microscopic studies were c onducted to determine the effect of acute and long-term (LTE) insulin exposure on insulin binding. Stock cultures included cells never expos ed to insulin and cultures grown in medium containing 6 mg/ml insulin. Logarithmic cultures were exposed to porcine insulin concentrations o f 0 and 6 mg/ml for 1 h (insulin treated, (IT)) after 0, 48 h, 1, 3, o r 6 months of LTE insulin exposure. 24 h after the 1 h insulin treatme nt, the cells were fixed, exposed to porcine insulin (antigen), proces sed immunocytochemically using a primary antibody to porcine insulin a nd a secondary antibody immunocytochemistry kit, and examined for stai ning intensity by video image analysis. Morphological observations con firm that T. pyriformis does bind insulin whether or not the cells hav e had prior exposure to insulin. IT increases insulin binding (up-regu lation) in previously unexposed cells (control, P<0.01) and produces a further amplification in cells having prior acute exposure (48 h) to insulin (P<0.01). However, LTE exposure to insulin (1, 3 and 6 months) caused a decrease in insulin binding (down-regulation) after IT (P<0. 01) such that LTE-IT cells were not different from control cells follo wing 1, 3 or 6 months of chronic insulin exposure to insulin. Staining intensity was not different between IT cells and cells cultured with insulin throughout the six month study. Results suggest that insulin b inding sites of T. pyriformis are subject to regulatory processes simi lar to those of metazoans.