Биологические науки / 8. Физиология человека и животных

Grigorova N.V., Psol M.A.

Zaporizhzhya National University

INFLUENCE OF PRELIMINARY STARVATION AND GLUCOSE INJECTION ON DEVELOPMENT OF ALLOXAN DIABETES IN GOLDEN HAMSTERS

Investigations of pancreatic islets state of animals under the condition of simulated diabetes mellitus is a very topical problem considering medical and social significance of this disease. Alloxan is referred to diabetogenic compounds which cause damage of insulin-producing cells in golden hamsters [1-3]. These data are confirmed with investigations based on usage of biochemical (determination of glucose level in blood) and functional (cytochemical reactions of aldehyde fuchsin and 8-(p-toluensulfonylamino)–quinoline (8-TSQ)) tests of pancreatic islets state [4]. It is of interest to study an influence of preliminary starvation (inhibitor of insulin secretion) and glucose injection (its specific stimulator) on state of pancreatic islets in golden hamsters who received alloxan.

For conducting this experiment we used 37 golden hamsters. 10 intact animals served as a control, others received injections of diabetogenic agent alloxan simultaneously with preliminary starvation and glucose load. Alloxan was injected subcutaneously in the dose of 200-400 mg/kg of 2-4% solution. In experiments with starvation animals were left without food for 24 hours. 40% solution of glucose was injected intraperitoneally in the dose of 10 g/kg.

For determination of glycemia with Haggedorn – Jensen method blood was taken from hamsters’ tails in 5 days after alloxan injection. Then golden hamsters were decapitated. Pieces of pancreas, extracted from animals, were fixed in Bowin’s liquid during 24 hours (for conducting of aldehyde fuchsin cytochemical reaction) and in cold acetone during 12 hours (for conducting of 8-TSQ cytochemical reaction).

Fixed in acetone pieces were driven to paraffins through xylols. In case of fixation in Bowin’s liquid pieces were preliminary carried through alcohols of rising concentrations, and then through xylols and paraffins.

For cytochemical revealing of zinc paraffinic sections of gland fixed in acetone were carried through xylols, alcohols, processed with acetone solution of 8-TSQ, rinsed in distilled water, embedded in glycerin, and examined in luminescent microscope (light filters V-1, Y-18). Yellow-green luminescent granules were revealed in cytoplasm of pancreatic beta cells on the slides. Intensity of luminescent reaction served as an indicator of zinc content in these cells.

For cytochemical determination of insulin paraffinic sections of gland fixed in Bowin’s liquid were carried through xylols, alcohols of declining concentrations, distilled water, then were stained with alcohol solution of aldehyde fuchsin, rinsed with muriatic alcohol, tap water, embedded in glycerin-gelatine, and examined in light microscope. Blue-purple granulation was revealed in cytoplasm of pancreatic beta cells on the slides. Abundance of granulation is an indicator of insulin contain in the cells.

Intensity of cytochemical reactions was evaluated with 3-point system which was introduced by Sokolovsky V.V. [5], Hayhoe F. and Quaglino D. [6]. One point was given for a weak positive reaction, two points for a moderate one, and three points for a highly intensive reaction.

For control group hamsters sugar level in blood was 6,4 + 0,21 mmol/l on average, and contains of zinc and insulin in pancreatic cells were 1,6 + 0,09 c.u. and 1,4 + 0,07 c.u. respectively. Under influence of alloxan glycemia increased on 167% (17,1 + 0,48 mmol/l; p < 0,001), and contains of zinc and insulin in insulin-producing cells dropped on 81% (0,3 + 0,04 c.u.; p < 0,001) and 86% (0,2 + 0,01 c.u.; p < 0,001) respectively.

Preliminary starvation of golden hamsters caused even further increase of glucose concentration in blood and decline of intracellular compounds. Glycemia amounted to 19,6 + 0,54 mmol/l, zinc concentration was 0,2 + 0,03 c.u. (p < 0,001), and contain of insulin in islets B-cells was 0,1 + 0,02 c.u. (p < 0,001), which correspond to 206% higher, 87% and 93% lower control. Prior glucose load compensate hyperglycemia, zinc and insulin deficiency in pancreatic islets. Under these circumstances level of sugar in blood was 13,7 + 0,35 mmol/l (p < 0,001), amount of metal was 0,9 + 0,06 c.u. (p < 0,001), and amount of hormone was 0,7 + 0,03 c.u. (p < 0,001) which correspond to 144% higher, 44% and 50% lower the standard.

Thus, alloxan cause development of hyperglycemia and lowering of zinc and insulin level in pancreatic cells B of golden hamsters. Animals, which were treated with alloxan under the condition of preliminary starvation, showed strongly pronounced alterations of investigated characteristics while prior glucose load cause less changes in these measures.

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