Insulin Withdrawal Research Articles

Circulating endothelin-1 levels in lean non-insulin-dependent diabetic patients: Influence of ACE inhibition

To evaluate the effect of captopril on plasma endothelin-1 (ET-1) levels and insulin sensitivity, 15 lean normotensive men (51.6 +/- 3.8 years) affected by non-insulin-dependent diabetes mellitus (NIDDM) underwent 2-h euglycemic hyperinsulinemic clamp. Each patient was then assigned to receive either captopril (25 mg twice daily for 1 week) or placebo, in a double-blind randomized fashion, before repeating clamp. At baseline, plasma ET-1 levels were 0.77 +/- 0.25 pg/mL in captopril (n = 10) and 0.83 +/- 0.3 pg/mL in placebo patients (n = 5). A twofold increase in plasma ET-1 levels occurred during the 2-h insulin infusion in both groups (P < .05 after 60 and 120 min), with a rapid return to baseline after 30 min from insulin withdrawal. After 1 week of therapy, total glucose uptake significantly increased in captopril (from 3.71 +/- 1.70 mg/kg/min to 4.24 +/- 1.72 mg/kg/min, P < .03) but not in placebo patients. Plasma ET-1 levels significantly decreased after captopril therapy (0.48 +/- 0.25 pg/mL at time 0, P < .03 v pretreatment levels), but were unaffected by placebo. Moreover, captopril slightly reduced the magnitude of ET-1 increment during insulin infusion (0.65 +/- 0.28 pg/mL and 0.88 +/- 0.48 pg/mL at 60 and 120 min, respectively, P < .05 v time 0). As a consequence, during the second insulin infusion circulating ET-1 levels were significantly lower in captopril- than in placebo-treated patients at time 0 (P < .02), 60 (P < .002), 120 (P < .004), and 150 min (P < .001).(ABSTRACT TRUNCATED AT 250 WORDS)

NIH3T3 Cells Transfected with the Yeast H+-ATPase Have Altered Rates of Protein Turnover

NIH3T3 cells transfected with the yeast plasma membrane H+-ATPase (RN1a line) or transfected with a low-activity mutant H+-ATPase (N-Mut line) were used to examine the relationship between cytosolic pH (pHcyt) and protein turnover. At an extracellular pH (pHex) of 7.15, NIH3T3 and N-Mut cells have a pHcyt of 7-7.1 and a vacuolar pH (pHvac) of 6.3, whereas in RN1a cells both the pHcyt and the pHvac are 0.3 unit more alkaline. Rates of protein synthesis and degradation are optimum at pHex 7.2 and are much more sensitive to pH changes in RN1a cells than in NIH3T3 cells. However, irrespective of pH, rates of protein degradation in RN1a cells are always less than those measured in NIH3T3 cells. Rates of protein synthesis are the same for sparse cultures of RN1a and NIH3T3 cells and show a density-dependent decline in NIH3T3 cells but remain high in RN1a cells even at high cell densities. These data indicate that the elevation of pHcyt caused by transformation with the H+-ATPase has no direct effect on protein synthesis. On the other hand, rates of protein degradation are consistently lower in RN1a cells than in NIH3T3 or N-Mut cells. Basal rates of protein degradation, measured in medium containing 10 mM 3-methyladenine or 10% serum or 1 μM insulin, as well as the autophagic response to serum or insulin withdrawal, are both significantly lower in RN1a cells. These data indicate that transformation with the H+-ATPase has a direct effect on rates of protein degradation, possibly through an elevation of pH. The higher pHvac will directly effect lysosomal protein breakdown and the higher pHcyt may be permissive for maintenance of low basal rates of protein breakdown. Overall, we conclude that transformation with the H+-ATPase provides a permissive environment for high rates of protein synthesis and low rates of protein degradation that result in high rates of growth and the tumor phenotype.

Modulation of P i transport in skeletal muscle by insulin and IGF-1

In vivo, skeletal muscle P i uptake influences both muscle cellular [P i] and plasma [P i], and may mediate the hypophosphataemic effects of insulin and insulin-like growth factor 1 (IGF-1). These effects were investigated in the cultured mouse myoblast cell line G8 and the isolated incubated rat soleus. The low K m for P i in G8 cells is consistent with in vivo evidence that muscle cell [P i] is partially protected against changes in plasma [P i]. Insulin and IGF-1 stimulated Na-dependent P i influx: in G8 cells both increased V max , with no change in K m , but while the insulin response occurred within 15 min and rapidly reversed upon insulin withdrawal, the response to IGF-1 occurred only after 60 min and persisted at least 60 min following IGF-1 withdrawal. Furthermore, only the IGF-1 response was inhibited by cycloheximide. We suggest that IGF-1 operates through de novo protein synthesis, while insulin stimulates transporter recruitment to the cell surface.

Effects of autonomic neuropathy on glucose, fatty acid, and ketone body metabolism following insulin withdrawal in patients with insulin-dependent diabetes

It has been postulated that sympathetic denervation due to autonomic neuropathy may protect against the development of diabetic ketoacidosis in patients with insulin-dependent diabetes. We tested this hypothesis by withdrawing exogenous insulin from five men with established insulin-dependent diabetes and severe symptomatic autonomic neuropathy. Six men also with insulin-dependent diabetes of more than 5-years duration but no clinical evidence of neuropathy served as controls. Normoglycemia was maintained during the night before the study using a variable intravenous insulin infusion that was terminated at 0800 h the following morning (time 0 min). During a 600-min observation period following termination of the insulin infusion the rise (mean ± SEM) in blood glucose concentration between 0 min and 600 min was comparable between the autonomic neuropathy and contol groups (9.2 ± 1.3 versus 8.3 ± 1.1 mmol/L; p > 0.1). The rise in plasma nonesterified fatty acids was significantly less for the autonomic neuropathy patients (0.96 ± 0.06 versus 1.65 ± 0.14 mmol/L; p < 0.01) with a similar trend for the rise in total ketone body concentration (2.74 ± 0.85 versus 3.72 ± 0.38 mmol/L). Capillary pH at 600 min, however, was not significantly different between the groups (23 ± 2 versus 20 ± 1 mmol/L). In conclusion, plasma fatty acids and ketone body concentrations rise less rapidly following withdrawal of insulin in patients with insulin-dependent diabetes and autonomic neuropathy suggesting that such patients may have a degree of protection against the development of diabetic ketoacidosis.

Insulin and okadaic acid induce Rab4 redistribution in adipocytes.

Insulin stimulation of glucose transport involves the translocation of vesicles containing the glucose transporter Glut 4 to the plasma membrane. Rab proteins, which have been implicated in the regulation of vesicular traffic, were studied in adipocytes. Rab3B, Rab3C, Rab4, and Rab8 were detected, but Rab3A was not. In the absence of insulin, Rab3B and Rab3C were cytosolic, while Rab4 and Rab8 were associated with membranes. Only Rab4 distribution was modified by insulin. In unstimulated adipocytes, most of Rab4 was found in a low density microsomal fraction, which also contained the majority of Glut 4. After insulin treatment, a 50% decrease in Rab4 content was observed, concomitantly with a departure of transporters to the plasma membrane. The dose responses for the departure of Glut 4 and Rab4 from the microsomal fractions were superimposable, half-maximal effects being obtained with 0.1 nM insulin. Rab4 was redistributed to the cytosol and its movement was reversed by insulin withdrawal. When Glut 4-containing vesicles were immunopurified with antibodies to Glut 4, Rab4 was found in the immune pellets, suggesting that Rab4 was tightly associated with the vesicles. Okadaic acid, an inhibitor of phosphatases 1 and 2A that is known to stimulate Glut 4 translocation, caused the same movement of Rab4 from low density microsomal fraction to the cytosol, while the phorbol ester 12-O-tetradecanoylphorbol-13-acetate had no effect. We suggest that insulin and okadaic acid induce a cycling of Rab4 from a vesicular fraction containing the Glut 4 transporter to the cytosol and that this cycling may participate in the insulin stimulatory action on glucose transporter translocation.

Short-term fasting is a mechanism for the development of euglycemic ketoacidosis during periods of insulin deficiency.

To determine the etiology of euglycemic ketoacidosis, the effect of a 32-h fast on the rate of metabolic deterioration was examined in a group of 10 healthy subjects with type I diabetes mellitus. Patients were studied during 5 h of insulin withdrawal after 8 h (postprandial) and 32 h (fasted) of food deprivation. Study parameters included substrate levels, electrolytes, counterregulatory hormone levels, and rates of glucose and glycerol turnover. In the fasted state, mean peak plasma glucose concentrations were significantly lower than those in the 8-h postprandial state (13.3 +/- 1.6 vs. 17.4 +/- 1.4 mmol/L, respectively; P < 0.05), and mean rates of glucose production were also significantly lower at all time points in the fasting state. The rate of development of ketosis was significantly more rapid during insulin deficiency after a fast (8.82 +/- 0.63 vs. 6.23 +/- 0.30 micro/L.min; P < 0.05), while plasma nonesterified fatty acids and glycerol turnover showed a biphasic response to insulin withdrawal, which was also more robust after a fast. Metabolic acidosis, as reflected in the rate of decrease in serum bicarbonate concentration, was more severe after 32 h of fasting than in the postprandial state (mean nadir, 15.4 +/- 0.9 vs. 18.6 +/- 0.5 mmol/L; P < 0.001). In contrast to values in the postprandial state, serum glucagon levels rose during insulin withdrawal in the fasting state, and plasma norepinephrine levels also correlated positively with the ongoing metabolic decompensation. Other counterregulatory hormones did not differ significantly in the fasted vs. postprandial states in these short term metabolic studies. We conclude that a fast of moderate duration, such as might be expected to occur during the development of diabetic ketoacidosis, predisposes patients with type I diabetes to euglycemic ketoacidosis during periods of insulin deficiency. Furthermore, decreased rates of hepatic glucose production are responsible for the lower plasma glucose values observed during a fast. The development of ketosis continued progressively in both conditions, but the rate of rise of plasma ketones was increased in the fasted state. This accelerated development of ketosis may be attributable to the effects of elevated levels of glucagon and/or catecholamines on lipolysis.

Glucose turnover in type I diabetic subjects during exercise. Effect of selective and nonselective beta-blockade and insulin withdrawal.

To assess the effect of selective beta 1-blockade (atenolol and betaxolol) and nonselective beta-blockade (propranolol) on glucose turnover in subjects with insulin-dependent (type I) diabetes mellitus during moderate exercise. Five subjects with type I diabetes were infused with insulin and then exercised for 1 h, after pretreatment with each of the three drugs or saline and, on a separate day, after withdrawal of insulin. Glucose turnover was measured using tritiated glucose. Plasma glucose, initially 9.2 +/- 0.5 mmol/L (mean +/- SE) when insulin infused and 14.0 +/- 0.8 when insulin was withdrawn, fell on exercise by 3.4 +/- 1.1 mmol/L (P < 0.05) saline, 4.0 +/- 0.8 mmol/L (P < 0.01) with betaxolol, 3.8 +/- 0.7 mmol/L (P < 0.01) with atenolol, 5.0 +/- 0.6 mmol/L (P < 0.005) with propranolol, and 1.7 +/- 1.0 mmol/L (NS) when insulin was withdrawn. Propranolol, but not the other beta-blockers, caused a significantly greater fall in glucose on exercise than during the control study. Glucose appearance rate (Ra) was similar basally and rose to an almost identical level in all five groups during exercise. Glucose disappearance rate (Rd) rose similarly during exercise, except after propranolol when the rise was significantly greater than with saline (P < 0.01). Failure of glucose to change significantly during exercise when insulin had been withdrawn was associated with the smallest rise in Rd and the highest nonesterified fatty acid concentrations. Propranolol and betaxolol, but not atenolol, reduced nonesterified fatty acids. We conclude that the greater fall in glucose on exercise after beta-blocking drugs is probably largely a direct effect of beta 2-blockade on muscle, increasing the exercise-induced rise in Rd glucose. This offers support to the use of beta 1-specific drugs, where beta-blockade is necessary in type I diabetes.

Transplantation of allogeneic fetal pancreases combined from MHC-different donor strains does not change rejection of the graft.

A number of 17.5- to 18.5-day-old fetal pancreases were grafted under the kidney capsule of streptozotocin-diabetic rats. Eight syngeneically grafted glands were sufficient to reverse the diabetes of the recipients within 4 weeks when the recipient rats were treated with insulin for 18 days after transplantation. Eight allogeneic fetal pancreases obtained from one donor strain were rejected after transplantation and the recipients relapsed into hyperglycemia immediately after insulin withdrawal. Eight allogeneic fetal pancreases obtained from eight MHC-different donor strains were also rejected and the recipients relapsed into hyperglycemia after insulin withdrawal. Using fetal pancreases as tissue sources, the combination of the allogeneic graft from different donor strains was not sufficient to prolong the survival time of the grafted tissue.

Transplantation of allogeneic fetal pancreases combined from MHC-different donor strains does not change rejection of the graft

Abstract. A number of 17.5-to 18.5-day-old fetal pancreases were grafted under the kidney capsule of strepto-zotocin-diabetic rats. Eight syngeneically grafted glands were sufficient to reverse the diabetes of the recipients within 4 weeks when the recipient rats were treated with insulin for 18 days after transplantation. Eight allogeneic fetal pancreases obtained from one donor strain were rejected after transplantation and the recipients relapsed into hyperglycemia immediately after insulin withdrawal. Eight allogeneic fetal pancreases obtained from eight MHC-different donor strains were also rejected and the recipients relapsed into hyperglycemia after insulin withdrawal. Using fetal pancreases as tissue sources, the combination of the allogeneic graft from different donor strains was not sufficient to prolong the survival time of the grafted tissue.

Differential regulation of multiple c- erbA expression by thyrotropin, insulin and insulin-like growth factor I in rat thyroid FRTL-5 cells

Regulation of multiple c- erbA gene expression was studied in rat thyroid FRTL-5 cells. Two species of erbA α (α1 and α2) mRNA and one species of erbA β (β1) mRNA were identified by Northern blot analysis. Withdrawal of thyrotropin (TSH), insulin and serum from the complete medium resulted in an increase in α1 and α2 erbA mRNA levels without altering the level of erbA β1 mRNA. Readdition of TSH, N 6,2′- O-dibutyryl cAMP or forskolin caused a transient reduction of α1 and α2 mRNA levels (75–90%) at 3–12 h. The α1 and α2 mRNA levels were restored at 24 h. The TSH action was dose-dependent showing the half-maximal effect at around 10 −9 M. Readdition of TSH did not show any effect on β1 mRNA level. The action of TSH was not dependent on ongoing protein synthesis but required ongoing transcription. Inhibitors of thyroid hormone biosynthesis, propylthiouracil and methylmercaptoimidazole, did not show any effect on TSH action. Readdition of insulin or insulin-like growth factor I (IGF-I) caused a dose-dependent reduction of α1 and α2 mRNA levels without any effect on β1 mRNA level. Their action was slower than TSH and persistent. The actions of insulin and IGF-I were dependent on both ongoing translation and transcription. These results indicate that TSH and insulin/IGF-I reduce levels of c- erbA α1 and α2 mRNA possibly by two distinct mechanisms without altering c- erbA β1 mRNA level in FRTL-5 cells.

Acute insulin withdrawal from diabetic BB rats decreases myocardial glycolysis during low-flow ischemia

We have previously demonstrated that withdrawal of insulin treatment from BB diabetic rats for a 24-hour period will increase the failure rate of hearts subjected to low-flow ischemia. The purpose of this study was to determine if this increased severity of ischemia was related to a decrease in glycolytic rates during ischemia. Two groups of insulin-dependent diabetic BB Wistar rats were used; in one group, insulin treatment was withheld from rats 24 hours prior to study (uncontrolled), while in the second group, the daily insulin injection was not withheld (insulin-treated). Isolated working hearts obtained from these animals were perfused with 30 mmol/L ( 2- 3H U- 14C )-glucose and 1.2 mmol/L palmitate, at an 11.5 mm Hg left atrial preload and 80 mm Hg aortic afterload. Hearts were subjected to a 15-minute aerobic perfusion followed by 60 minutes of low-flow ischemia (coronary flow, 0.5 mL/min). Under aerobic conditions, steady-state glucose oxidation rates (measured as 14CO 2 production) were decreased in the uncontrolled group compared with the insulin-treated group (85.3 ± 21.5 v 406.2 ± 120.1 nmol/min/g dry weight, respectively; P < .05). Steady-state glycolytic rates (measured as 3H 2O production) were also decreased in the uncontrolled group compared with the insulin-treated group (1.73 ± 0.30 v 5.57 ± 1.26 μmol/min/g dry weight, respectively; P < .05). During low-flow ischemia, glucose-oxidation rates markedly decreased in both groups (23.9 ± 8.7 and 38.3 ± 25.2 nmol/min/g dry weight in the uncontrolled and insulin-treated diabetic rats, respectively). Glycolytic rates during ischemia decreased slightly, but remained significantly depressed in the uncontrolled group compared with the insulin-treated group (1.28 ± 0.28 v 2.15 ± 0.25 μmol/min/g dry weight, respectively; P < .05). During reperfusion, glucose oxidation recovered to preischemic values in hearts from uncontrolled diabetic rats, and remained depressed compared with the insulin-treated diabetic rats (84.7 ± 9.2 and 366.9 ± 81.3 nmol/min/g dry weight, respectively, P < 0.05). Glycolytic rates also returned to preischemic values and remained depressed in these hearts (2.27 ± 0.42 and 4.71 ± 0.91 μmol/min/g dry weight, respectively, P < .05). Combined with our previous studies, these data suggest that poor control of insulin-dependent diabetic BB rats increases the sensitivity of the heart to ischemia, possibly by decreasing the ability of hearts to maintain high glycolytic rates during ischemia.

Effect of pancreas transplantation on glucose counterregulation in insulin‐dependent diabetic patients prone to severe hypoglycaemia

Pancreatic transplantation was performed in three patients with insulin-dependent diabetes mellitus in whom recurrent and severe episodes of hypoglycaemia had been found to be due to defective glucose counterregulation. Thus in these patients the spontaneous blood glucose recovery after insulin-induced hypoglycaemia (0.1 U kg-1 h-1 i.v. insulin until blood glucose levels fell below 2.8 mmol l-1) was delayed, and the responses of glucagon, epinephrine and growth hormone (GH) were absent or diminished. After pancreas transplantation the patients exhibited essentially normal blood glucose control. When the insulin infusion test was repeated 3 months after the transplantation, the blood glucose level recovered rapidly after insulin withdrawal. The glucagon response was restored, and the responses of epinephrine and GH were improved. Plasma C-peptide was suppressed by approximately 50%, which is less than is observed in normal subjects. It is concluded that glucose counterregulation improves after pancreas transplantation. This appears to be mainly due to an improvement in the hypoglycaemia-induced glucagon response, but an amelioration of sympatho-adrenal and hypothalamic-pituitary regulatory mechanisms may also be involved. The apparent failure to suppress completely the insulin release from the denervated pancreas transplant indicates that inhibition of beta-cell secretion during insulin-induced hypoglycaemia may be partly under neural control.

Preventive effects of octreotide (SMS 201‐995) on diabetic ketogenesis during insulin withdrawal.

1. Exogenous somatostatin inhibits glucagon secretion and prevents ketoacidosis in diabetic patients, but has the therapeutic disadvantage of requiring continuous intravenous infusion to exhibit these effects. 2. Consequently, we examined the effect of subcutaneous administration of the long-acting somatostatin analogue octreotide (SMS 201-995) on early ketogenesis in diabetic ketoacidosis. On two separate occasions insulin was withdrawn over a period of 9 h from seven type I diabetic patients. On the second occasion the patients were given 50 micrograms octreotide s.c. before the insulin withdrawal and every 3 h during insulin withdrawal. 3. Differences in integrated free fatty acid responses (4706 +/- 1227 mumol l-1 h vs 3026 +/- 835 mumol l-1 h, AUC, P = NS) were not significant, but the peak increments of acetoacetate (1413 +/- 354 mumol l-1 vs 612 +/- 176 mumol l-1, P less than 0.05), beta-hydroxybutyrate (2180 +/- 475 mumol l-1 vs 922 +/- 246 mumol l-1, P less than 0.01) and the decrements in plasma bicarbonate (-8 +/- 1 mumol l-1 vs -4 +/- 1 mumol l-1, P less than 0.05) and pH (-0.07 +/- 0.01 vs -0.03 +/- 0.01, P less than 0.05) were significantly less with octreotide. 4. At the same time peak increments of glucagon were lower with octreotide treatment (329 +/- 206 pg ml-1 vs 39 +/- 30 pg ml-1, P less than 0.05). 5. We conclude that, despite accelerated lipolysis and provision of substrate for ketogenesis during insulin withdrawal, this somatostatin analogue significantly reduces ketogenesis resulting from insulin deprivation, probably secondary to decreasing glucagon secretion. This drug may be useful in short term prophylactic treatment of diabetic patients during periods of increased risk for ketoacidosis.

Differential effects of insulin deficiency on albumin and fibrinogen synthesis in humans.

Insulin deficiency decreases tissue protein synthesis, albumin mRNA concentration, and albumin synthesis in rats. In contrast, insulin deficiency does not change, or, paradoxically, increases estimates of whole body protein synthesis in humans. To determine if such estimates of whole body protein synthesis could obscure potential differential effects of insulin on the synthetic rates of individual proteins, we determined whole body protein synthesis and albumin and fibrinogen fractional synthetic rates using 5-h simultaneous infusions of [14C]leucine and [13C]bicarbonate, in six type 1 diabetics during a continuous i.v. insulin infusion (to maintain euglycemia) and after short-term insulin withdrawal (12 +/- 2 h). Insulin withdrawal increased (P less than 0.03) whole body proteolysis by approximately 35% and leucine oxidation by approximately 100%, but did not change 13CO2 recovery from NaH13CO3 or estimates of whole body protein synthesis (P = 0.21). Insulin deficiency was associated with a 29% decrease (P less than 0.03) in the albumin fractional synthetic rate but a 50% increase (P less than 0.03) in that of fibrinogen. These data provide strong evidence that albumin synthesis in humans is an insulin-sensitive process, a conclusion consistent with observations in rats. The increase in fibrinogen synthesis during insulin deficiency most likely reflects an acute phase protein response due to metabolic stress. These data suggest that the absence of changes in whole body protein synthesis after insulin withdrawal is the result of the summation of differential effects of insulin deficiency on the synthesis of specific body proteins.

Expression of growth hormone, growth hormone releasing hormone, and somatostatin genes is unperturbed in the streptozotocin-induced diabetic rat

The effects of streptozotocin diabetes on the level of growth hormone, growth hormone releasing hormone, and somatostatin mRNA was measured in control rats, in diabetic rats maintained on insulin, and in diabetic rats in which insulin had been withheld for 3 days. Total cytoplasmic RNA samples were prepared from the pituitary and hypothalamic tissues of each animal and analyzed by dot blot or Northern blot hybridization. No significant difference was observed between control and insulin-treated groups with regard to body weight or plasma glucose concentration. The insulin withdrawal group had significantly higher plasma glucose concentrations and lower body weights, confirming diabetic status. There was no significant difference in the level of growth hormone, growth hormone releasing hormone, and somatostatin mRNA among any of the three groups however. We conclude that alterations in the regulation of circulating growth hormone in the streptozotocin-induced diabetic rat, removed from insulin treatment for 3 days, did not occur at the transcriptional or RNA processing level. This conclusion extends to hypothalamic growth hormone releasing hormone, and somatostatin gene expression as well. Regulatory changes in growth hormone level previously noted during insulin withdrawal in the streptozotocin-induced diabetic rat could be the result of post-transcriptional processes operating at the level of hormone synthesis or release.

The effect of insulin withdrawal on intermediary metabolism in patients with diabetes secondary to chronic pancreatitis

Insulin was withdrawn from 7 patients with Type I (insulin-dependent) diabetes and 4 patients with insulin-dependent diabetes secondary to chronic pancreatitis, both groups without residual beta-cell function. Median plasma glucagon concentrations rose slightly, but significantly after withdrawal of insulin in Type I diabetic patients (from 14 (range: 11-16) to 19 (14-25) pmol/l by 6 h), but not in the patients with secondary diabetes. This was accompanied by a significantly higher increase in blood glucose concentration from 5.1 (4.9-5.7) to 15.2 (12.9-18.1) mmol/l by 6 h in Type I diabetic patients compared with patients with secondary diabetes (from 4.9 (4.3-6.7) to 13.1 (10.9-13.5) mmol/l) (p less than 0.01). Beta-hydroxybutyrate increased to a similar extent in the two groups, whereas no significant increases were found in glycerol and lactate in any of the groups. Increased secretion of glucagon is not essential for the development of hyperglycemia and ketonemia in patients with diabetes secondary to chronic pancreatitis, but may augment the degree of hyperglycemia in Type I diabetic patients compared with patients having secondary diabetes.

Measurement of whole-body protein turnover in insulin-dependent (type 1) diabetic patients during insulin withdrawal and infusion: comparison of [13C]leucine and [2H5]phenylalanine methodologies

1. The aims of this study were twofold: (i) to investigate the ability of a recently described [2H5]phenylalanine method for quantifying whole-body protein turnover during acute physiological perturbation; (ii) to determine specifically whether the previously observed increase in protein synthesis on insulin withdrawal in insulin-dependent (type 1) diabetic patients seen when employing the [13C]leucine technique could be corroborated by using [2H5]phenylalanine. 2. Whole-body protein turnover was measured by both the [2H5]phenylalanine and [13C]leucine primed continuous infusion methods applied simultaneously to six type I post-absorptive diabetic patients during insulin withdrawal and infusion. 3. Values were determined by the [13C]leucine method by measuring either [13C]leucine (primary pool) or alpha-[13C] ketoisocaproic acid (reciprocal pool) enrichment in plasma. 4. Values of whole-body protein breakdown during insulin withdrawal derived from the [2H5]phenylalanine and primary and reciprocal pool [13C]leucine models respectively were 3.54 +/- 0.43, 3.85 +/- 0.41 and 4.62 +/- 0.44 g day-1 kg-1 (means +/- SD). Insulin infusion resulted in a significant reduction (P less than 0.02) to 3.07 +/- 0.34, 3.05 +/- 0.26 and 3.82 +/- 0.4 g day-1 kg-1, respectively. Synthesis values fell significantly but by a smaller amount than breakdown, resulting in increased (P less than 0.05) net protein deposition, regardless of the model used. 5. These data demonstrate that the [2H5]phenylalanine and [13C]leucine methods generate similar results both in absolute and relative terms in response to short-term insulin infusion. 6. The confirmation of increased whole-body protein synthesis during insulin withdrawal by two independent methods supports the validity of this observation.

Plasma Catecholamine Responses to Hypoglycemia in Children and Adolescents With IDDM

The goal of this study was to assess whether children and adolescents with insulin-dependent diabetes mellitus (IDDM) have decreased catecholamine responses to insulin-induced hypoglycemia as has been reported in adults and to explore the pathogenesis of the decreased response in terms of possible relationships to autonomic neuropathy or hyperinsulinism. A before-and-after trial on the effects of 3 days of intensive insulin therapy was conducted on 60 subjects with IDDM (age 15.4 +/- 2.6 yr, duration of diabetes 7.8 +/- 3.5 yr). The control group consisted of 5 children with non-growth hormone-deficient short stature (age 14.8 +/- 3.2 yr). Hypoglycemia was induced with an intravenous insulin bolus (0.15-0.75 U/kg) after insulin withdrawal and 3 days of intensive insulin therapy in diabetic subjects on an inpatient basis to assess the role of hyperinsulinism in suppressing the catecholamine response to hypoglycemia. Control subjects were studied once and received an insulin bolus of 0.1 microU/kg. Autonomic neuropathy was assessed by computerized assessment of the basal R-R variation during inspiration and expiration and the pancreatic polypeptide response to hypoglycemia. Basal plasma catecholamine levels were lower in diabetic subjects after intensive insulin therapy than in control subjects (P = 0.008). The peak and incremental catecholamine responses after insulin withdrawal and intensive insulin therapy in IDDM subjects were significantly decreased compared with control subjects (P less than 0.001). Peak catecholamine responses to hypoglycemia in IDDM were decreased after intensive insulin therapy (P = 0.002). This was particularly true in those with plasma glucose nadir levels of less than 2.2 mmol (P less than 0.001). The diminished catecholamine responses were primarily due to decreased peak epinephrine responses after intensive insulin therapy compared with insulin withdrawal (P = 0.011). There were no significant correlations between the catecholamine response to hypoglycemia and age, duration of diabetes, pancreatic polypeptide, or R-R interval. These results suggest that children and adolescents with IDDM after insulin withdrawal have diminished catecholamine response to hypoglycemia compared with control subjects and indicate that short-term intensive insulin therapy diminishes this response further. Thus, hyperinsulinism may play a role in suppressing the catecholamine response to hypoglycemia. There is no evidence for a clinical or subclinical role of autonomic neuropathy to explain the altered catecholamine responses.

Altered behavior and neurochemistry during short-term insulin withdrawal in streptozocin-induced diabetic rats.

Amphetamine-induced stereotypy and turnover of dopamine (DA) and serotonin (5-HT) in the CNS were assessed in streptozocin-induced diabetic rats 48 h after withdrawal from insulin treatment and compared with nondiabetic controls, diabetic rats receiving continued insulin treatment, and chronically hyperglycemic diabetic rats. Stereotypy was attenuated in chronically hyperglycemic diabetic rats but normalized in insulin-treated diabetic rats. Stereotypy in insulin-withdrawn rats was intermediate, being attenuated significantly relative to controls but not significantly different from either insulin-treated diabetic rats or chronically hyperglycemic diabetic rats. DA turnover was significantly reduced in striatum and hypothalamus in all diabetic groups. 5-HT turnover was reduced in chronically hyperglycemic diabetic rats in all four brain regions but normalized in insulin-treated diabetic rats. Insulin-withdrawn diabetic rats had significantly reduced 5-HT turnover in frontal cortex. Taken together, the findings indicate that insulin replacement does not normalize diabetes-induced reduction in DA turnover and that short-term insulin withdrawal, e.g., occurring clinically with noncompliance, affects both 5-HT turnover in some brain regions and behavior.

Altered behavior and neurochemistry during short-term insulin withdrawal in streptozocin-induced diabetic rats

Altered behavior and neurochemistry during short-term insulin withdrawal in streptozocin-induced diabetic rats