Growth hormone injected daily in 6 dogs for 6 days caused a 20-fold elevation in fasting serum immunoreactive insulin (IRI) without appreciable change in serum glucose in 1 day. In the somatotrophic diabetes that occurred after 2 days, the hyperinsulinaemia was maintained and the serum IRI/glucose (I/G) ratio declined from the early high level but remained elevated. During this treatment, in response to glucose infusion, the rise in serum IRI above the initially high fasting level was 16 times the normal. In response to glucagon, the rise in IRI was twice the normal and the rise in glucose was more prolonged, resulting in a decline in the I/G ratio. In response to arginine infusion, the rise in serum IRI was 8 times the normal and the rise in the I/G ratio was twice normal. Following a meal, the rise in serum IRI was 8 times the normal. Thus, with growth hormone treatment the insulin secretory responses to these stimulating factors were magnified over the already elevated fasting level of secretion. The insulin content of the pancreas was reduced to less than 10% of normal by growth hormone treatment for 6 days, due apparently to elevation of the rate of secretion over the rae of formation of insulin.

The relation of insulinase activity to the biological effect of insulin on isolated tissue was studied. Rat diaphragm in vitro caused the rapid disappearance of immunoreactive insulin (IRI) in physiological concentrations. IRI loss at time intervals was exponential. The fractional rate of loss of IRI was therefore independent of IRI concentration and was also approximately constant per milligram of tissue, the value being 0.0216%/mg∙mm. The value of the Michaelis constant (Km), obtained from initial velocities at five initial concentrations of IRI, was 1.85 × 10−8M, and of the maximal velocity (Vmax) was 2.32 × 10−11 mole/g∙min, based on insulin dimer. The addition of an insulinase inhibitor (a partial hydrolysate of insulin) to hemidiaphragm in vitro reduced the fractional rate of IRI loss by 60%. The increase in Km, without appreciable change in Vmax, indicated that the inhibition was competitive. The IRI preserved by means of the inhibitor was biologically active, since it increased the glycogen accumulation and the incorporation of 14C-U-glucose into glycogen in a second, fresh hemidiaphragm. In single incubation of hemidiaphragm with insulin, the gain in glycogen was correlated with the amount of inhibitor. The biological effect of insulin on diaphragm in vitro was therefore limited by tissue insulinase activity, and insulinase inhibitor potentiated to some extent the action of insulin.