Leonard L. Madison

Studies of muscle capillary basement membranes in normal subjects, diabetic, and prediabetic patients

A technique is described for the measurement of muscle capillary basement membranes by electron microscopic examination of needle biopsies of the quadriceps muscle. With this procedure it has been possible to obtain an objective evaluation of the significance of capillary basement membrane hypertrophy in diabetic microangiopathy. The results of such studies of muscle capillary basement membrane thickness in 50 normal, 51 diabetic, and 30 prediabetic patients have demonstrated the following. First, that the average capillary basement membrane width of diabetic patients is over twice that of normal subjects; moreover, such basement membrane thickening is a very constant finding among overtly diabetic patients, in that approximately 98% of individual diabetic subjects demonstrated this lesion. The degree of basement membrane thickening in diabetic patients is, however, unrelated to age, weight, severity, or duration of diabetes. Second, capillary basement membrane hypertrophy has been found in approximately 50% of patients who are genetically prediabetic but who have not yet demonstrated evidence of the manifest carbohydrate disturbances of diabetes mellitus. Third, in contrast to the results obtained in genetically diabetic patients, subjects with severe hyperglycemia due to causes other than genetic diabetes only infrequently show basement membrane hypertrophy. These results indicate that thickening of the muscle capillary basement membranes is a characteristic of genetic diabetes mellitus, and further, that the hyperglycemia of diabetes is probably not the factor responsible for the microangiopathy characteristic of diabetes mellitus. Finally, the discovery of thickened capillary basement membranes in prediabetic patients suggests that basement membrane hypertrophy is a relatively early lesion of the diabetic syndrome and provides further support for the conclusion that this vascular defect is independent of carbohydrate derangements of diabetes mellitus.

Physiologic Effects of Metabolic Fuels on Carbohydrate Metabolism: I. Acute Effect of Elevation of Plasma Free Fatty Acids on Hepatic Glucose Output, Peripheral Glucose Utilization, Serum Insulin, and Plasma Glucagon Levels

The in vivo effects of acute changes in the concentration of plasma free fatty acids on hepatic glucose output and peripheral glucose utilization were defined in thirteen studies in dogs. Dogs with chronic end-to-side portacaval shunts . were used thereby permitting not only measurement of hepatic rather than splanchnic glucose output but also the rate of peripheral glucose utilization. In eighteen additional studies the effect of acute elevations of plasma free fatty acids on serum insulin, plasma glucagon, blood ketones and alpha-amino nitrogen was determined. The quantitative and qualitative changes in plasma free fatty acids which occur during starvation were simulated by the intravenous administration of a cottonseed oil emulsion plus heparin. The acute elevation of plasma free fatty acids produced a 37 per cent decrease in hepatic glucose output and a 30 per cent inhibition of peripheral glucose utilization. This decrease in hepatic glucose output was the consequence both of a significant increase in the secretion of insulin and a decrease in glucagon secretion. These changes in hormonal milieu which attended the increase in plasma free fatty acids could not be ascribed to changes in blood ketones or amino acids since the concentrations of these substances remained unchanged. These data suggest that this feedback of free fatty acids on secretion of insulin and glucagon plays an important physiologic role during starvation in the safe transition from carbohydrate to fat metabolism without the danger of progressive ketoacidosis. it is likely that during starvation plasma free fatty acids contribute to the control and maintenance of a low but vital concentration of plasma insulin.

Glucagon antibodies and their use for immunoassay for glucagon.

Summary Non-precipitating antibodies to glucagon have been produced in rabbits and detected by both Berson-Yalow and Skom-Talmadge technics. A basis for an immunoassay for glucagon appears, thereby, to have been established.

Effect of plasma free fatty acids on plasma glucagon and serum insulin concentrations

Abstract The acute elevation of plasma free fatty acids to levels found during prolonged starvation by means of infusion of a triglyceride emulsion plus heparin in 5 dogs was associated with a 40 per cent decrease in plasma glucagon and a 33 per cent increase in serum insulin. This inhibitory feedback of free fatty acid on glucagon secretion and stimulatory feedback on insulin secretion very likely contribute to the smooth transition from carbohydrate to fat metabolism characteristic of prolonged starvation without the danger of progressive ketoacidosis.

Ethanol-induced hypoglycemia: I. The acute effects of ethanol on hepatic glucose output and peripheral glucose utilization in fasted dogs☆

Abstract Sixteen experiments were performed on dogs with chronic end-to-side portacaval shunts fasted 2 to 3 days to determine the acute effects of ethanol on hepatic glucose output and peripheral glucose utilization. Dogs with chronic end-to-side portacaval shunts were used since this preparation not only permits measurement of hepatic rather than splanchnic glucose balance but also allows calculation of peripheral glucose utilization. In 8 additional studies the effects of ethanol on splanchnic glucose output was determined in nonshunted healthy mongrel dogs fasted a similar period of time. Ethanol administered at mean rates varying from 0.029 to 0.183 mM/Kg./min. produced both a prompt (65 per cent) fall in mean hepatic glucose output and 25 per cent inhibition of peripheral glucose utilization. Hypoglycemia eventuated only when the magnitude of decrease in hepatic glucose output exceeded the magnitude of inhibition of peripheral glucose utilization. Splanchnic glucose balance during ethanol infusion also decreased significantly in dogs fasted for 2 to 3 days. These data show that ethanol-induced hypoglycemia is entirely the consequence of a decrease in the hepatic release of glucose. Its occurrence in glycogen-depleted starved dogs supports the hypothesis that ethanol inhibits hepatic gluconeogenesis, decreases hepatic release of glucose and thereby produces hypoglycemia. Since ethanol not only reduces hepatic glucose output but also inhibits peripheral glucose utilization, the failure to produce arterial hypoglycemia cannot be construed as evidence that ethanol does not consistently alter carbohydrate metabolism in fasted animals.

Ethanol-induced Hypoglycemia: II. Mechanism of Suppression of Hepatic Gluconeogenesis

The postulation that the increased NADH2/NAD ratio generated in the liver cell during ethanol metabolism causes the suppression of hepatic gluconeogenesis has been tested in several ways in twenty-eight fasted glycogendepleted dogs in whom hepatic gluconeogenesis was inhibited by infusions of ethanol. First, it was shown that fructose, a non-NAD-dependent precursor of glucose, produced a rapid restoration of hepatic glucose output during ethanol-induced suppression of hepatic gluconeogenesis. Second, in contrast, the infusion of glutamate and α-ketoglutarate, both NAD-dependent precursors of glucose, failed to augment the depressed rate of hepatic gluconeogenesis induced by ethanol. Finally, the administration of methylene blue, a redox dye which oxidizes NADH2 to NAD, not only prevented the expected fall in hepatic glucose output when infused simultaneously with ethanol, but also produced a rapid restoration of hepatic glucose output previously depressed by ethanol administration in fasting dogs. These data are consonant with the thesis that the increased NADH2/NAD ratio, which characterizes ethanol oxidation by the liver cell, causes a partial block at several points in the gluconeo genic pathway and is responsible for the ethanol-induced suppression of hepatic gluconeogenesis.

Abnormal Alpha Cell Function in Diabetics Response to Insulin

The extremely high levels of glucagon recently observed in dogs with severe alloxan-induced diabetes decline promptly and precipitously to normal as soon as exogenous insulin is infused. This suggests that the normal response of the pancreatic alpha cell to hyperglycemia requires the presence of circulating insulin. To determine if the relative hyperglucagonemia of human diabetics responds similarly to insulin repletion, the plasma glucagon response of ten adult-type diabetic patients to a large, predominantly carbohydrate meal was determined with and without the simultaneous forty-five-minute intravenous infusion of glucagonfree insulin (0.12 to 0.2 U./kg.). The glucagon response to the carbohydrate meal during prompt and supernormal hyperinsulinemia resulting from the infusion did not differ from that of the control meal, i.e. normal suppression of glucagon by hyperglycemia was not restored by the abundance of circulating insulin. To determine if still higher plasma levels of insulin would overcome the hyposuppressibility of the diabetic alpha cell to hyperglycemia, 0.6 U. per kilogram per hour of insulin was infused at a constant rate for two hours together with 0.6 gm. per kilogram per hour of glucose to prevent hypoglycemia. Insulin levels of more than 1,200 μ,U. per milliliter were thus attained. Under these conditions, plasma glucagon declined from a mean preinfusion level of 97 pg./ml. (SEM ± 11) to a nadir of 75 pg./ml. (SEM ± 10) ninety minutes later. This slow, modest, statistically significant (p < 0.01) decline differed strikingly from the response of eight nondiabetic patients given intravenous glucose alone; in these subjects, at a comparable level of hyperglycemia, glucagon declined from a mean fasting level of 90 pg./ml. (SEM ± 8) to 57 pg./ml. (SEM ± 8) within thirty minutes, despite an insulin rise to only 46 μU./ml. It was concluded that in human diabetics the acute restoration of plasma insulin, even to supernormal levels, does not promptly restore to normal the alpha cell responsiveness to hyperglycemia. Simple insulin lack may not, therefore, adequately explain the alpha cell abnormality in human diabetes.

The physiological significance of the secretion of endogenous insulin into the portal circulation. IV. Hepatic uptake of glucose during glucose infusion in non-diabetic dogs.

Recent studies from this laboratory (1) have shown that insulin has an immediate and profound effect upon net hepatic glucose balance. When insulin was administered at a rate (2) estimated to minimize the counter-regulatory responses to hypoglycemia, the decline in blood glucose which ensued was largely the consequence of a diminished hepatic release of glucose (1). Since, under physiologic circumstances, endogenous insulin secretion is stimulated by the rising blood glucose concentration (3, 4) which follows a carbohydrate meal, the effect of insulin in regulating hepatic glucose metabolism is best assessed in the presence of a glucose load. Whether or not hyperglycemia, with its attendant release of endogenous insulin, results in a decrease or cessation of hepatic glucose output has been a controversial subject (5-13). However, in all but one of the previous studies (5), the conclusions were based upon inferential data, since net hepatic glucose balance was not measured. The present studies were undertaken to determine the effect of hyperglycemia and the concomitant release of endogenous insulin upon hepatic glucose metabolism. Hyperglycemia was produced by the intravenous infusion of glucose. The rate of glucose administration was varied in order to define better some of the factors that con-

Effect of phenoformin on peripheral glucose utilization in human diabetic and nondiabetic subjects.

Neither the precise biochemical mechanisms whereby phenethylbiguanide (DBI) lowers blood glucose concentration in human subjects nor its site of action, whether in the peripheral tissues and/or the liver, are known. In the absence of a urinary loss of glucose or glucose intermediaries, hypoglycemia can result only from an increase in peripheral glucose utilization or a decrease in hepatic output of glucose. Although many factors may be involved in either increased peripheral utilization or decreased hepatic production of glucose, the localization of the site of action of DBI might offer some insight into its possible mechanism of action. The present study was designed to determine the effect of DBI on peripheral glucose utilization in diabetic and normal human subjects. Changes in peripheral glucose utilization* were measured by following the changes in femoral arteriovenous glucose concentration difference after the administration of DBI.

Primary thrombosis of the internal carotid artery: Report of seven cases with cerebral circulatory and metabolic studies

Abstract The literature on spontaneous internal carotid artery thrombosis is reviewed and seven additional cases are reported. The clinical characteristics of the disorder are discussed, with special reference to those features which are of particular importance with respect to clinical recognition. Cerebral circulatory studies made in these seven patients did not reveal hemodynamic or metabolic changes that could be convincingly attributed to unilateral internal carotid artery thrombosis alone. However, the studies of others 30 have shown that interruption of the flow of blood through one internal carotid artery is accompanied by an increase in CVR. Our results indicate that neurologic sequelae in internal carotid artery thrombosis occur only when this increment in CVR is superimposed upon a pre-existing increase in CVR which is of sufficient magnitude to reduce CBF to a level which is insufficient to maintain the structural integrity of the entire brain. They emphasize the fact that it is the state of the CVR prior to internal carotid artery occlusion that determines whether neurologic complications will occur. It is suggested that cerebral circulatory studies should be done, whenever feasible, before therapeutic carotid artery ligation, for they provide important and otherwise unobtainable information regarding the safety of the procedure. In one of the patients an attempt was made to revascularize the brain by creating an artificial common carotid artery-internal jugular vein fistula on the side of the thrombosed internal carotid artery. Postoperative cerebral angiograms and studies of arterial-internal jugular venous oxygen difference showed this procedure to be ineffective.