Summary. The effects of chronic elevations of growth hormone levels on hepatic insulin binding and action were studied in rats with subcutaneously implanted growth hormoneproducing tumours. These animals were significantly heavier (p < 0.001 ; 388 _+ 29 versus 239_+ 4 g), had elevated insulin levels, (6.8 _ 0.6 versus 3.3 _ 0.5 ng/ml), lower glucose concentrations (6.0_+ 0.4 versus 9.3 _ 0.5 mmol/1) and larger hepatocyte diameters (28.7_ 0.6 versus 24.0_+ 0.4 p~m) and surface areas (2661 + 119 versus 1835 + 65 am 2) than control rats. Insulin binding and action [net (C~4)-glucose incorporation into glycogen] were compared in hepatocytes isolated from these two groups. Because of the difference in hepatocyte size, insulin binding was normalized for cell surface area. Binding of the tracer alone (0.9 _+ 0.05 versus 1.3 _+ 0.12%/cm:) and capacity of the high affinity, low capacity receptor (30.9_+ 5.9 versus 65.2 _+ 5.9 sites/Ixm 2) were significantly decreased (p < 0.02) in tumour-bearing rats. Dose-response curves of insulin action in hepatocytes chronically exposed to excess growth hormone were shifted to the fight. The maximal response was also significantly decreased. However, the relation between the amount of insulin bound and the proportion of the maximum insulin effect obtained were similar in cells from the two groups. Thus, in rat hepatocytes chronically exposed to excess growth hormone, both a receptor and a post-receptor defect occur while the insulin receptor itself functions normally.
Material and MethodsGH3 cells were supplied by the American Type Culture Collection (Bethesda, Maryland). These cells are an established clonal strain of rat pituitary tumour cells secreting both GH and prolactin [12]. Cells were maintained in monolayer culture in a humidified atmosphere of 95% air -5% CO2 using Dulbecco's modified Eagle's medium (DMEM) supplemented with horse serum (15%), fetal calf serum (2.5%), penicillin (50 U/ml), streptomycin (50 p~g/ml) and glutamine (5 mmol/1) (Irvine Scientific, Santa Ana, California). For tumour generation, approximately 5-7 • 106 cells derived from the same parent culture were harvested by trypsinization, washed and resuspended in DMEM containing penicillin (50 U/ml) and streptomycin (50 p~g/ml). An aliquot of the cell suspension was counted in an automatic cell counter (Coulter Electronics, Hialeah, Florida), and the suspension was diluted to -106 cells/ml. One ml was injected SC into female Wistar-Furth rats as described previously [11]. Tumours were palpable by the end of 4 weeks. Control rats were treated similarly except that they received the media alone. Tumour-bearing and control rats were sacrificed alternately 6-8 weeks after inoculation.Rats were anaesthetized with sodium pentobarbital (60 mg/kg). A cannula was placed in the portal vein to prepare isolated hepatocytes (see below) but before the perfusion was started, blood was sampled from the inferior vena cava via a small needle attached to a syringe. Glucose concentrations were measured by a glucose oxidase met...
