The effect of space flight on the board of the satellite Cosmos 2044 on plasma hormone levels and liver enzyme activities of rats.

The aim of present experiment was to study the changes of corticosterone, insulin and glucose levels in plasma, of the activity of enzymes involved in aminoacid metabolism in liver and the binding of insulin to specific receptors of cell membrane from liver and also of adipose tissue of rats exposed to space flight for 14 days on biosatellite Cosmos 2044. Adult male Wistar rats (body mass 300-370 g) were divided into five groups: intact control rats (AC), rats exposed to space flight (F), animals in synchronous model experiment (S), rats in antiorthostatic hypokinesia (A) and so called operated control group (C). Half of all groups (5 animals) except the intact control were operated 3 days before the experiment (fibulas on both hind legs were broken). The flight animals were sacrificed 5-6 hours after landing. It was observed that plasma insulin levels are increased in rat exposed to 14-day space flight and in synchron experiments. A significant increase of plasma glucose levels was found in flight rats in spite of high insulin concentrations suggesting that in rats exposed to 14-day space a deterioration of tissue sensitivity to insulin could by present. No significant differences of specific insulin binding to liver plasma membrane fraction in flight and intact control animals were observed. A decrease of insulin binding capacity in liver was found in rats in antiorthostatic hypokinesia (A). However in the membrane of adipocytes an important increase of insulin receptors was noted in rats subjected to space flight. These results suggest, that the liver and adipocyte insulin receptors of flight rats did not respond to the increased plasma insulin levels by "down regulation". The determination of plasma corticosterone levels showed that in flight rats and animals exposed to antiorthostatic hypokinesia the plasma hormone levels are significantly elevated. A significant increase of tyrosine aminotransferase and tryptophan pyrrolase activities in liver of flight rats and those exposed to hypokinesia was observed. Also the elevation of alanine amino-transferase in liver was observed in flight rats, while, the activity of aspartate aminotransferase in liver was similar in control and flight animals. These results showed that the changes in liver enzyme activities in rats after 14-day space flight are in agreement with the results observed in previous experiments after a shorter space flight (7 days).

Comparison of radiochemical purity and tissue binding of labelled insulin prepared by lactoperoxidase and chloramine T iodination.

The radiochemical purity and tissue binding of 125I-insulin labelled by conventional chloramine T method were compared with those of (A14)-monoiodoinsulin prepared by lactoperoxidase iodination. In the first case the labelled insulin was purified on a cellulose column, while (A14)-monoiodoinsulin prepared with the aid of lactoperoxidase was purified on a column of QAE Sephadex A-25. After the digestion of labelled insulin by pronase about 15 percent of diiodoinsulin was found after chloramine T iodination, while a negligible amount (i.e. less than 1 percent) was detected after the use of lactoperoxidase. In addition, no damage of insulin molecules was found after the use of the latter method and the purification on QAE Sephadex A-25 yielded a homogenous preparation of insulin. Specific binding to isolated rat fat cells, rat liver plasma membranes and human erythrocytes was consistently higher in the case of (A14)-monoiodoinsulin compared to the insulin labelled by chloramine T method.

Effect of n-hexane inhalation on insulin degradation in rat blood in vitro.

Effect of inhalation of an air-n-hexane mixture on insulin degradation in rat blood in vitro was studied under simultaneous monitoring of n-hexane concentrations in blood (using gas chromatography) and in the inhaled air (using infrared analysis). Inhalation of n-hexane vapours in a concentration of 54,694 +/- 1933 ppm (197 +/- 7 g m-3) resulted in blood n-hexane saturation within 15 min. (0.18 +/- 0.023 mg ml-1) associated with a statistically increased (P less than 0.002) insulin degradation in the blood of exposed rats as compared to control animals 0.152 +/- 0.03 vs. 0.053 +/- 0.003). To rule out possible effects of stress on the results obtained, effects of both, 20 min restraint stress (RS) and equally long handling (HAN) on insulin degradation were also estimated. The data thus obtained were compared with insulin degradation in control (C) rats which were killed by decapitation in the animal room. No significant differences were observed between the individual groups (C: 0.055 +/- 0.08; RS: 0.068 +/- 0.01; HAN: 0.069 +/- 0.015). In vitro addition of n-hexane to pure plasma or to a mixture of blood elements revealed that degradation activities present in the cellular blood compartment are required for the n-hexane-induced enhancement of insulin degradation in the blood to occur.

Changes of insulin in plasma and receptor for insulin in various tissues after the exposure of rats to space flights and hypokinesia.

The explanation of the mechanism of the response to gravity changes is of great importance for the determination of the capacity of human subjects to adapt to the load of gravitational stress. Therefore several studies were performed to investigate the activity of endocrine system, since the hormones are involved in the regulation of physiological functions and metabolic processes. However the studies of endocrine system activity during altered gravity conditions, especially during the weightlessness are influenced by the several interventions in biomedical observations due to operational program of astronauts, wide variability in individual response and tolerance, use of extensive countermeasures, differences in the type of space missions and in the studies after landing also a hypergravity effect at landing and variability in postflight readaptation process. The significant changes of plasma insulin and glucose levels were observed in astronauts during space flights and in the first days of recovery period. In the first inflight period plasma insulin levels were increased, unchanged or decreased however after 4-5 weeks of exposure to weightlessness a decrease of insulin plasma levels were noted. After space flights an increase of plasma insulin levels were demonstrated in experimental animals and in human subjects. Since plasma insulin level is considered as most important factor involved in the regulation for insulin receptors in target tissues, an investigation of insulin receptors in various tissues was performed in rats exposed to space flight or to hypokinesia (model used for simulation of some effects of microgravity).

Acute stress stimulates the red cell sodium pump in rat.

The effect of restraint stress (RS) on the red cell sodium pump, hematocrit, plasma osmolality and plasma sodium and potassium concentrations was measured. Ten min of RS did not produce any change in the sodium pump activity. Longer lasting stress - 150 min - stimulated significantly the sodium pump activity in comparison to the control group (8.52 +/- 0.76 vs. 7.78 +/- 0.46 min-1 per 10(13) Ery; P less than 0.05; means +/- S.D.). Hematocrit rose during the stress (controls: 44.8 +/- 2.3; 10 min RS: 48.1 +/- 1.1; 150 RS: 47.0 +/- 1.9%; P less than 0.01 and P less than 0.05, respectively). The values for osmolality and plasma sodium concentration did not change. However, the plasma potassium concentration decreased significantly after 150 min of RS (4.63 +/- +/- 0.74 vs. 3.3 +/- 0.48 mmol 1(-1); P less than 0.001). Thus, acute stress stimulates the sodium pump activity in erythrocytes of the rat and norepinephrine seems most likely to trigger this activation.

Restriction fragment length polymorphism of the insulin receptor gene, type 2 diabetes and insulin binding.

In searching for a genetic marker of type 2 diabetes we estimated the frequency of alleles of the Bgl II restriction fragment length polymorphism (RFLP) of the insulin receptor gene in a group of type II diabetic patients (n = 50), characterized by OGTT (glucose, insulin, C-peptide) and insulin receptor binding parameters. Leucocyte DNA was incubated with restriction endonuclease Bgl II and specific fragments were determined by Southern blot technique, using radioactive plasmid pINSR 13.1 as insulin receptor gene probe for hybridization. Insulin receptor numbers and receptor affinity were estimated by 125I-(Tyr-A-14)- insulin binding to red blood cells. Among control subjects the 20 kb fragment (allele Bgl II+) had a frequency of 0.21. In our group of diabetic patients this allele had a frequency of 0.10 (n.s., p greater than 0.05). In our study the insulin receptor genotype had no influence on body mass index, insulin and C-peptide during OGTT as well as insulin receptor binding data. So far, etiopathogenetic linkage between diabetes and insulin receptor variants (mutants) could unambiguously be proved in patients with extreme insulin resistance only. In our opinion, the estimation of the role of the gene as the reason underlying the disease inevitably requires the investigation of large families with multiple occurrence of type 2 diabetes.