Abstract

A recently developed indigestible dextrin (IDex) was studied for its effects on glucose tolerance in male Sprague–Dawley rats. IDex is a low viscosity, water-soluble dietary fibre obtained by heating and enzyme treatment of potato starch. It has an average molecular weight of 1600.

An oral glucose tolerance test was conducted with 8-week-old rats to evaluate the effects of IDex on the increase in plasma glucose and insulin levels after a single administration of various sugars (1·5 g/kg body weight). The increase in both plasma glucose and insulin levels following sucrose, maltose and maltodextrin loading was significantly reduced by IDex (0·15 g/kg body weight). This effect was not noted following glucose, high fructose syrup and lactose loading.

To evaluate the effects of continual IDex ingestion on glucose tolerance, 5-week-old rats were kept for 8 weeks on a stock diet, a high sucrose diet or an IDex-supplemented high sucrose diet. An oral glucose (1·5 g/kg body weight) tolerance test was conducted in week 8. Increases in both plasma glucose and insulin levels following glucose loading were higher in the rats given a high sucrose diet than in the rats fed a stock diet. However, when IDex was included in the high sucrose diet, the impairment of glucose tolerance was alleviated. Moreover, IDex feeding also significantly reduced accumulation of body fat, regardless of changes in body weight.

These findings suggest that IDex not only improves glucose tolerance following sucrose, maltose and maltodextrin loading but also stops progressive decrease in glucose tolerance by preventing a high sucrose diet from causing obesity.

Journal of Endocrinology (1995) 144, 533–538

ABSTRACT

Two groups of monolayer cultures of pancreatic cells from the neonatal rat were maintained in glucosedepleted TCM 199 medium, supplemented with 5·5 mmol galactose/l, with or without 0·1 mmol 2-deoxyglucose/1. Another group was kept in medium with 5·5 mmol galactose/l alone, following exposure for 2 days to a medium with 5·5 mmol galactose/1 and 10 μmol iodoacetic acid/l to kill fibroblasts selectively. Each of these monolayers was cultured in a perifusion system for a total of 7 days so that phasic insulin secretion could be compared. On day 0, B cells responded in a monophasic fashion to acute challenge with 16·7 mmol glucose/l whereas, in the presence of 10 μmol forskolin/1 and 1 mmol 3-isobutyl-1-methyl-xanthine/l, the same dose of glucose stimulated a biphasic response of approximately the same magnitude. At a concentration of 10 mmol/l, leucine and 2-ketoisocaproate both produced only minimal increases in the second phase of secretion above the basal level. No response to secretagogues was seen under culture conditions without 2-deoxyglucose. In contrast, addition of 2-deoxyglucose to the galactose-supplemented medium stimulated B cells to secrete insulin in a biphasic fashion in response to a single dose of glucose, and the stimulatory effects of leucine and 2-ketoisocaproate were also remarkably increased. Moreover, when exposed to a linear concentration gradient of glucose, leucine or 2-ketoisocaproate, these B cells responded to secretagogues in a dose-dependent fashion. On the other hand, B cells which had been exposed briefly to iodoacetic acid and then cultured in medium with galactose alone, showed a biphasic response to glucose, leucine and 2-ketoisocaproate; however, quantitative relationships differed. Thus the response (total insulin secreted during a 30-min stimulation) of B cells grown in 2-deoxyglucose to glucose was 610%, to leucine 504% and to 2-ketoisocaproate 810% of that of cells exposed to iodoacetic acid. The present data suggest that cultivation of neonatal B cells in a glucose-depleted medium containing 2-deoxyglucose may cause functional maturation.

J. Endocr. (1987) 115,169–175

ABSTRACT

The effect of two concentrations of glucose on the maturation of the response of B cells was studied in pancreatic monolayer cultures of the neonatal rat using a perifusion system. After exposure for the initial 3 days to a medium with 5·5 mmol glucose/l and 10 μmol iodoacetic acid/l (day 3), in order to delete fibroblasts selectively, monolayer cultures were kept for a total of 12 days in medium with either 5·5 or 16·7 mmol glucose/l alone. On day 3, B cells responded in a monophasic fashion, with no significant second phase, to acute challenge with 16·7 mmol glucose/1. At a concentration of 10 mmol/l, leucine and 2-ketoisocaproate both produced only minimal increases in the second phase of secretion above the basal level. In contrast, B cells on day 7 cultured in 5·5 mmol glucose/l showed a biphasic response to glucose, leucine and 2-ketoisocaproate. The magnitude in response to glucose was well preserved at day 15 of culture, whereas the stimulatory effects of leucine and 2-ketoisocaproate decreased to 24–57% of that of B cells on day 7. Moreover, B cells on day 7 cultured in 16·7 mmol glucose/l responded in a biphasic manner to glucose, the response being 65% of that of B cells in 5·5 mmol glucose/l. Additionally, the response to leucine and 2-ketoisocaproate still appeared to be monophasic. At day 15 of culture, however, the response of B cells in 16·7 mmol glucose/l to glucose was 105%, to leucine 245% and to 2-ketoisocaproate 127% of that of B cells in 5·5 mmol glucose/l. In conclusion, these results suggest that cultivation in medium with 5·5 mmol glucose/l could induce precocious development of an adult pattern of insulin secretion compared with that in 16·7 mmol glucose/l and, in addition, that the medium with 16·7 mmol glucose/l is superior for long-term culture to the medium with 5·5 mmol glucose/l.

J. Endocr. (1988) 118, 303–310

ABSTRACT

The effect of maternal hyperglycaemia on the function of neonatal B cells was examined using a perifusion technique in pancreatic monolayer cultures of neonatal rats from normoglycaemic mothers (C), and those made slightly hyperglycaemic (SH) and highly hyperglycaemic (HH) by injection of streptozotocin. Monolayer cultures were kept for 7 days in medium containing 5·5 mmol glucose/l plus 1 mmol 2-deoxy-glucose/l. On day 0, B cells in the C group responded to 16·7 mmol glucose/l, 10 mmol leucine/l and 10 mmol 2-ketoisocaproate/l in a monophasic fashion with no significant rise in the second phase. However, compared with the C group, a significant increase in the second-phase secretion in response to glucose and 2-ketoisocaproate was observed in the SH group, although there was no difference in the first-phase secretion. In the HH group the insulin secretion was lower in the first phase but not in the second phase. After culture for 7 days, B cells in the C group showed a biphasic response to the secretagogues, with a great increase in the second-phase secretion. In the SH group, the second phase of insulin secretion was increased but the increment was far less than that in the C group. The secretory response was remarkably low in the HH group compared with other groups. From these results, we conclude that at an early stage of culture slight maternal hyperglycaemia causes a hypersensitivity of neonatal B cells but impairs the normal development of the function of B cells during culture, and that high hyperglycaemia results in impaired insulin secretion throughout the whole period of culture studied.

J. Endocr. (1988) 119, 493–499

ABSTRACT

Techniques for the monolayer culture of pancreatic islet cells from adult rats and the responsiveness of B cells are described. Whole pancreatic tissue was enzymatically dispersed and then cultured for 30 days in tissue culture medium 199 containing 5·5 mmol glucose/l, with or without 1 mmol 2-deoxyglucose/l. In the absence of 2-deoxyglucose, the responsiveness of B cells diminished to almost zero by day 15 and islets degenerated. In contrast, addition of 2-deoxyglucose to the medium resulted in a selective degeneration of fibroblasts, yielding monolayers that consisted mostly of islet cells. In this stationary system in which monolayers of islet cells were maintained in medium with 2-deoxyglucose, insulin secretion from B cells on days 15 and 30 increased in a dose-dependent fashion in response to increasing concentrations of glucose, leucine and 2-ketoisocaproate. Similarly, when exposed to 16·7 mmol glucose/l, perifused B cells showed a biphasic pattern of insulin secretion on day 15. Addition of 10 μmol forskolin/l and 200 nmol 12-O-tetradecanoyl phorbol13-acetate/l remarkably enhanced this response. Likewise, the response to 10 mmol leucine/l or 10 mmol 2-ketoisocaproate/l was biphasic. These results suggest that these monolayer cultures retain the functional properties of the adult rat pancreas, and may be useful not only as a model for the in-vitro study of B cell function, but also for implantation.

J. Endocr. (1988) 118, 173–178

ABSTRACT

Monolayer cultures of the pancreas of the neonatal rat were maintained for 7 days in glucose-depleted TCM 199 medium, supplemented with 5·5 mmol galactose/l, with or without 0·1 mmol 2-deoxyglucose/l. Under culture conditions without 2-deoxyglucose, the responsiveness of B cells supported on galactose was totally abolished by day 7 of culture, and islets degenerated. In contrast, the addition of 2-deoxyglucose to the galactose-supplemented medium promoted the survival and the function of B cells even in a glucose-depleted environment and, in addition, yielded the monolayers mostly consisting of endocrine cells by destroying fibroblasts selectively. On day 7 the recovery of insulin in the cells was higher than that of the cells grown in medium with galactose alone (10·7-fold), and than the initial level at day 0 (twofold). Furthermore, the response to an acute challenge with 16·7 mmol glucose/1 was 3·3-fold, to 10 mmol leucine/1 it was 8·5-fold, and to 10 mmol 2-ketoisocaproate/l it was 10·9-fold above each value observed on day 1. In summary, the above data indicate that morphologically intact, functionally competent endocrine B cells can be grown in medium free from glucose.

J. Endocr. (1984) 103, 377–381

Abstract

The synaptic membrane protein synaptosomal-associated protein (SNAP-25) has recently been implicated as one of the key proteins involved in exocytotic membrane fusion in neurons. However, the role of SNAP-25 in pituitary hormone release is not known. In this study, we determined that SNAP-25 is involved in regulated exocytosis in the clonal pituitary cell line GH4C1. SNAP-25 messenger RNA and protein were detected in GH4C1 cells by RT-PCR and immunoblot analysis, respectively. Immunofluorescence analysis indicated that SNAP-25 protein was localized in the plasma membrane. Next, to determine the function of SNAP-25 in GH4C1 cells, specific inhibitors of SNAP-25, botulinum neurotoxin (BoNT) /A or /E, and antisense SNAP-25 oligonucleotide were used. Neither BoNT/A nor BoNT/E affected thyrotropin-releasing hormone (TRH)-induced cytosolic Ca²⁺ increase, but both inhibited TRH-induced exocytosis. Moreover, they dose-dependently inhibited TRH-induced prolactin release. The introduction of antisense oligonucleotide into the cells also inhibited TRH-induced prolactin release. These results suggest that SNAP-25 is involved in regulated exocytosis in GH4C1 cells.