Bernard W. Ince

The effects of pancreatic hormones, catecholamines, and glucose loading on blood metabolites in the northern pike (Esox lucius L.)

Abstract A serial blood sampling technique involving heart cannulation was used to study the effects of bovine insulin and glucagon, codfish insulin, adrenalin, noradrenalin, and glucose loading on blood glucose, plasma amino acid nitrogen, and cholesterol in the northern pike, Esox lucius . Bovine insulin at 10, 25 and 50 IU/kg produced hypoglycaemia and death 24–72 hr after injection. At 2 IU/kg, however, a transient hypoglycaemia was observed with return to normal levels after 24 hr. Codfish insulin at 2 IU/kg produced hypoglycaemia which persisted for up to 7 days. Bovine and codfish insulin lowered plasma amino acid nitrogen significantly, although return to normality was observed only at 2 IU/kg. Neither codfish nor bovine insulin alone altered plasma cholesterol levels. Glucagon at 1 mg/kg and 2 mg/kg produced hyperglycaemia at 0.5 hr which persisted for 9 hr before return to normal levels. Glucagon had no effect on either plasma amino acid nitrogen or cholesterol. Adrenalin at 0.05 mg/kg, 1 mg/kg, and 5 mg/kg produced a significant hyperglycaemia at 0.5 hr, a return to normal levels after 6 hr, and a progressive hypoglycaemia thereafter. Adrenalin at the above dose rates was generally without effect on either plasma amino acid nitrogen or cholesterol, although at 5 mg/kg, a progressive lowering of cholesterol was observed. Noradrenalin at 1 mg/kg resulted in hyperglycaemia followed by a significant hypoglycaemia after 24 hr. It was without effect on either plasma amino acid nitrogen or cholesterol. A single glucose load of 0.5 g/kg resulted in an increase in blood glucose followed by a slow return to normal levels after 48 hr, whilst the levels of amino acid nitrogen and cholesterol were unaffected. Neither the shape of the glucose-loading curve nor the levels of metabolites were significantly altered by 4 weeks of starvation. Bovine insulin (10 IU/kg) and glucose (0.5 g/kg), when injected simultaneously, resulted in a greatly improved glucose tolerance with a reduction, and subsequent recovery, of plasma amino acid nitrogen. Levels of plasma cholesterol were significantly lowered 48 hr after a combined insulin and glucose injection. These results suggest that insulin and glucagon control the circulating levels of glucose in the blood. In addition to glucagon, the catecholamines are also able to effect a rapid glycogenolysis, possibly providing for increased glucose supplies during periods of predatory activity. The influence of insulin on amino acid nitrogen in the pike may reflect an important role in this and other wholly carnivorous forms.

Plasma insulin levels in teleosts determined by a charcoal-separation radioimmunoassay technique.

Abstract Plasma insulin levels in a variety of teleosts were determined by a charcoal-separation radioimmunoassay technique employing cod insulin components. The method was satisfactory for all species studied but was more sensitive for cod and less so for trout, dab, and eel, while the values for plaice and pike were at the lower limits of sensitivity of the assay. Plasma insulin and glucose levels in cannulated European silver eels, sampled over 4 days, were both elevated initially but subsequently fell to stable, lower values after 2 days. Plasma insulin and amino-acid nitrogen levels of fed rainbow trout and cod were significantly higher than for starved animals while the correlation of plasma insulin with glucose was less apparent. The study illustrates the need for careful consideration of the blood-sampling technique to be used and of the nutritional state of the fish whose plasma insulin is to be measured.

Effects of insulin and of metabolite loading on blood metabolites in the European silver eel, (Anguilla anguilla L.)

Abstract The effects of codfish and bovine insulin, and of glucose and amino acid loading on blood glucose, plasma cholesterol and amino acid nitrogen were studied in the European silver eel, Anguilla anguilla . Serial blood samples were withdrawn with minimal stress from individual animals after cannulation of the bulbus arteriosus. The operation initially caused marked elevations of the metabolites, necessitating a 48 hr post-operative recovery period. Codfish insulin at 2 IU/kg produced a significant yet recoverable lowering of blood glucose, plasma cholesterol and amino acid nitrogen, while bovine insulin at the same does level effected a reduction in amino acid nitrogen only. A single intra-arterial glucose load of 0.5 g/kg resulted in an increase in blood glucose levels followed by a slow return to normal after 24 hr. Plasma cholesterol levels were significantly lowered after such a treatment while amino acid nitrogen remained unaltered. A repeated intra-arterial glucose load 24 hr later resulted in an enhanced rate of glucose removal from the blood. A glucose load administered via the intestine resulted in a smaller increase in blood glucose, with more rapid return to normal levels, while plasma cholesterol and amino acid nitrogen remained unaltered. An intra-arterially injected mixture of amino acids (0.25 g/kg) was rapidly removed from the blood within 3 hr. In addition, plasma cholesterol levels were reduced and a rapid hyperglycaemia developed, followed by hypoglycaemia 9 hr later. An intestinal load of amino acids resulted in a slower rate of removal from the blood together with a sustained hyperglycaemia, but of lesser degree than after intra-arterial administration. Plasma cholesterol levels remained unaltered by an intestinal amino acid load. The results indicate a greater sensitivity of the European silver eel to codfish rather than bovine insulin, and point to the possibility of an important role of insulin in the metabolic control of the initial phases of its spawning migration. The metabolite-loading data indicate that mechanisms for the removal of excess glucose and amino acids are still functional in pre-migratory silver eels and may be partly insulin-dependent.

Amino acid stimulation of insulin secretion from the in situ perfused eel pancreas; Modification by somatostatin, adrenaline, and theophylline

Amino acid stimulation of insulin secretion, and its modification by somatostatin, adrenaline and theophylline, has been studied using the in situ perfused eel pancreas. Lysine, leucine, and phenylalanine at 10 mM caused biphasic insulin release, while histidine produced a weak, nonphasic response by comparison. Lysine was the most effective secretagogue in this series. Somatostatin (10 and 100 ng/ml) and adrenaline (10 and 50 ng/ml) significantly reduced total insulin secretion in response to 10 mM lysine. At the lower concentration the hormones were without effect on first-phase insulin secretion but significantly lowered second phase secretion. At the higher concentration, however, first phase secretion was significantly reduced, and second phase secretion virtually abolished. Theophylline (2 mM), increased basal insulin secretion in a biphasic manner and significantly potentiated 10 mM lysine-stimulated insulin secretion. These data indicate an important role of somatostatin and adrenaline in the control of B-cell function in teleosts and further, suggest that insulin-secretory responses, as in mammals, may be cyclic AMP dependent.

Plasma insulin and glucose responses to glucagon and catecholamines in the European silver eel (Anguilla anguilla L.)

Plasma insulin and glucose responses to intra-arterial injections of glucagon, adrenalin, and noradrenalin were studied in cannulated European silver eels (Anguilla anguilla L.). Glucagon (50, 100, and 200 μg/kg) was without effect on plasma insulin, whereas both adrenalin (25 and 50 μg/kg) and noradrenalin (50 and 100 μg/kg) caused a biphasic response, an initial significant depression over 30 min to 1 hr, followed by a significant elevation after 3 hr. Repeated injections of noradrenalin (100 μg/kg) inhibited the elevation of plasma insulin observed 3 hr following a single injection, and insulin release caused by arginine (25 mg/kg) was also inhibited by a simultaneous injection of adrenalin (50 μg/kg). The hormones caused significant hyperglycaemia at all dose levels, adrenalin being the most potent. It is suggested that catecholamine-induced inhibition of insulin release may have some physiological significance in permitting rapid mobilisation of metabolic fuels during periods of high activity.

Hormonal and metabolite effects on plasma free fatty acids in the northern pike, Esox lucius L

Abstract The effects of intraarterial injections of bovine and codfish insulin, glucagon, adrenalin, noradrenalin, glucose, and amino acids on plasma free fatty acid (FFA) levels were studied in the Northern pike, Esox lucius L., which were cannulated to permit serial blood sampling. Codfish insulin caused a prolonged reduction of FFA while bovine insulin (2 IU/kg) was without effect. Glucagon (2 and 4 mg/kg) significantly increased FFA between 6 and 9 hr. Adrenalin (0.05 and 1 mg/kg) lowered FFA below control levels between 1 and 3 hr. Noradrenalin (0.05 mg/kg) was without effect on FFA whereas at 1 mg/kg an initial increase and subsequent decrease was observed. An amino acid mixture (0.25 g/kg) lowered FFA while glucose (0.05 g/kg) was without effect. The opposing effects of insulin and glucagon on the plasma FFA point to their possible regulatory functions in the lipid metabolism of the pike. The lowering of FFA by amino acids suggests an insulin-assisted response which is not seen following glucose injection. The inability of adrenalin to stimulate lipolysis in the pike contrasts sharply with its well-known effects in mammals.

The in vivo metabolism of 14C-glucose and 14C-glycine in insulin-treated northern pike (Esox lucius L.).

Abstract The effects of codfish insulin (2 IU/kg) on 14C-glucose and 14C-glycine metabolism in vivo were studied in cannulated Northern pike (Esox lucius L.). Insulin significantly reduced plasma glucose levels and increased the incorporation of 14C-glucose carbon into liver and muscle lipid and muscle protein, and of 14C-glycine carbon into muscle protein. These results suggest that the metabolic role of insulin in pike is similar to that observed in mammals.

Insulin kinetics and distribution in rainbow trout (Salmo gairdneri)

The plasma disappearance of intravenously-injected 125I-cod insulin and a mixture of 131I-beef insulin and 125I-beef monoiodoinsulin, studied in rainbow trout (Salmo gairdneri), was found to be multiexponential, exhibiting both a rapid and a slow component. Biological half-disappearance times (T14) and metabolic clearance rates (MCR) varied significantly with three different assay methods for 125I-cod insulin, while the “apparent” distribution spaces (DS) were similar for all methods. The immunoassay technique with anti-fish insulin serum provided a significantly greater discrimination between total radioactivity and 125I-cod insulin radioactivity than either precipitation with 20% trichloracetic acid (TCA) or charcoal adsorption. The labelled beef insulins were assayed by double-antibody immunoprecipitation using anti-beef insulin serum. The MCR for 125I-beef monoiodoinsulin (1.78 ± 0.20 ml/kg/min) was significantly greater than for either 131I-beef insulin (0.58 ± 0.06 ml/kg/min) or immunoassayable 125I-cod insulin (0.74 ± 0.06 ml/kg/min). The results support the current view that monoiodoinsulin is a more valid tracer for insulin kinetic studies than randomly iodinated forms. A tissue distribution study of 125I-cod insulin showed the presence of TCA-precipitable radioactivity in all tissues and fluids studied, but the kidney was found to be the major accumulation site.