Tissue specific differences in insulin receptor characteristics in the mature bovine animal

Abstract

Insulin is believed to play a key role in the partitioning of nutrients towards fat and protein deposition. The biological effects of insulin are a function of plasma insulin concentration, insulin-receptor concentration and the affinity of the receptor. The number and affinity of insulin receptors on adipocytes differs between genetically lean and obese pigs (Camara and Mourot, 1996) while receptor number, but not affinity, was shown to differ in a variety of bovine muscles (Boge et al., 1995). The objective of the current work was to determine if insulin receptor affinity and number vary between the principal target tissues (i.e. liver, muscle and adipose tissue) in the bovine animal.Five Charolais-cross steers were offered grass silage ad libitum from 18 months of age until slaughter, at 701± 22.9 kg, at a commercial abattoir. Samples of liver [L], of two skeletal muscles [Ml, M3] and of subcutaneous [S], omental [0]and renal [R] fats were taken as soon as possible after slaughter (typically <30min for L, M3, S and O and <45min for Mland R), wrapped and snap frozen in liquid nitrogen prior to storage at -70 °. Insulin receptors were partially purified by solubilising tissues in 1% Triton X-100 for 24 hours at 4 °C followed by centrifugation (100000 x g, lh, 4°) and affinity chromatography of the supernatants on 0.5 ml wheatgerm agglutinin Sepharose 6MB columns. Bound receptors were eluted from the column with 0.3M N-acetyl-D-glucosamine. Receptor binding was assessed using a tracer amount of A-14 125I-insulin (∼35 pM) and increasing concentrations of unlabelled insulin (0-10 μM) in a total volume of 150 μl of pH 7.4 buffer as described by Magri et al (1990).