Effects of some 1,4‐dihydropyridine Ca antagonists on the blast transformation of rat spleen lymphocytes

Ca antagonists of different classes (verapamil, nifedipine, nicardipine, diltiazem) in a concentration of 10⁻⁵ m and higher are known to suppress Ca²⁺ transport into the lymphocyte cytosol, changing a normal response of lymphocytes to mitogens and antigens and so inhibiting their proliferation, as well as IL‐2‐induced cell proliferation, and their receptor expression on the surface of lymphocytes without cell cytotoxicity. In the present work we studied the effect of some 1,4‐dihydropyridines (DHP) such as nimodipine, nicardipine, nifedipine, niludipine, cerebrocrast, etaftoron, as well as metabolites of cerebrocrast: compounds 7 and 8, (four of the last were synthesized in the Latvian Institute of Organic Synthesis) on rat spleen isolated lymphocyte activation and proliferation in vitro following stimulation with the mitogens concanavalin A (Con A) and recombinant interleukin‐2 (IL‐2), insulin and insulin antibodies. Based on the experimental results we conclude that in low concentrations (10⁻⁷ to 10⁻⁹ M ) the tested 1,4‐DHP Ca antagonists stimulated the process of rat spleen lymphocyte proliferation and DNA synthesis, especially cerebrocrast. It is proposed that these Ca antagonists, as well as causing a concentration decrease of Ca²⁺, also activated phosphodiesterase, which in its turn, suppressed cAMP accumulation in the lymphocytes and eventually increased Ca²⁺ ion transport in the cells. Cerebrocrast among all the studied DHP Ca antagonists was the most potent in studies of activation of the lymphocytes in the presence of Con A, IL‐2 and insulin, which indicates the number of suppressor and helper lymphocytes and formation of insulin and interleukin receptors on their membrane surface. The increase in the lymphocyte suppressive activity produced by this compound effect can prevent diabetes mellitus types I and II at the stages of pre‐diabetes, early and distant diabetes, from hyperexpression of insulin and its receptor antibodies. Copyright © 1999 John Wiley & Sons, Ltd.     

Effect of cerebrocrast on the lymphocyte blast transformation activity in normal and streptozotocin‐induced diabetic rats

Both IDDM and NIDDM are characterized by deviations in peripheral T and B lymphocyte count, T_helper:T_suppressor ratio, as well as by impaired T_suppressor function. These abnormalities may promote insulin antibody and other antibody production, contributing to overt diabetes mellitus development in early stage of the disease. In the present study we explored the effects of cerebrocrast (1,4‐dihydropyridine derivative) administration on Con A‐ and IL‐2‐stimulated tissue lymphocyte blast transformation activity and on the thymus and lymph node mass in normal and streptozotocin (STZ)‐induced diabetic rats. It was established that cerebrocrast, administered four times at the doses of 0·05 and 0·5 mg kg⁻¹, has long‐term (up to 14 days) effects on the immune system and protects against the toxic effect of STZ in STZ‐induced diabetic rats, preventing thymus and lymph node mass loss. We conclude that cerebrocrast administration leads to the increase in number and activity of T_helper and T_suppressor lymphocytes. Glycolysis and DNA synthesis in these cells is augmented under the influence of cerebrocrast administration. We propose that the increase in lymphocyte suppressive activity caused by cerebrocrast administration may prevent the development of IDDM and NIDDM in patients with pre‐diabetes, but in patients with early and overt diabetes mellitus the drug administration may prevent the overexpression of insulin antibodies and other antibodies. The effect of cerebrocrast on the de novo production of insulin and IL‐2 receptors may be beneficial for IDDM and NIDDM patients. Copyright © 1999 John Wiley & Sons, Ltd.     

Protective effect of cerebrocrast on rat brain ischaemia induced by occlusion of both common carotid arteries

Diabetes mellitus is accompanied by several cardiovascular complications including atherosclerosis, cerebral ischaemia and stroke. We examined the neuroprotective effect of a 1,4-dihydropyridine derivative cerebrocrast (C, a new antidiabetic agent, synthesized in the Latvian Institute of Organic Synthesis) on the level of ATP in the brain, and on changes of the EEG and ECG, as well as blood pressure parameters in anaesthetized Wistar male rats before and during 10-min occlusion of both common carotid arteries. Cerebrocrast was administered i.v. at doses of 1.0 and 10 microg/kg in the v. femoralis 20 min prior to ischaemia. After 10-min ischaemia animals were decapitated and the brain was immediately frozen in liquid nitrogen and subsequently used for analysis of changes of ATP contention. Cerebrocrast, administered at doses of 1.0 and 10 microg/kg 20 min prior to occlusion of both common carotid arteries, completely prevented a fall in the ATP content of brain compared with the control rats. In control rats the content of ATP in brain during ischaemia decreased from 2.77 +/- 0.22 (basal level) to 1.74 +/- 0.20 micromol/g as a result of ischaemia. By administration of cerebrocrast 20 min before occlusion of the arteries, the content of ATP in the brain remained at the level of preischaemia (1.0 microg/kg C + ischaemia 2.82 +/- 0.36; 10 microg/kg C + ischaemia 2.42 +/- 0.22 micromol/g). Analysis of EEG parameters both before and during 10 min of occlusion showed that at a C dose of 1.0 microg/kg before occlusion produced a regular alpha rhythm during ischaemia and prevented cerebral bioelectric activity from significant changes. The depression of basal rhythm was observed at a C dose of 10 microg/kg during ischaemia in two rats out of six as well as an increase in the ECG ST segment above the isoelectric line. Blood pressure was decreased by about 10-20 mm Hg. We propose that pretreatment of rats with cerebrocrast at doses of 1.0 or 10 microg/kg 20 min prior to ischaemia can prevent ischaemic damage of rat brain, maintain necessary energy consumption, promote ATP production in brain cells, and prevent significant changes in EEG and ECG parameters. These properties are important in diabetes mellitus and its evoked cardiovascular complications as stroke, ischaemia, etc.     

Effect of new and known 1,4-dihydropyridine derivatives on blood glucose levels in normal and streptozotocin-induced diabetic rats

Analysis of the effect of several 1,4-DHP Ca(2+) channel antagonists on experimental and clinical diabetes shows that structurally similar Ca(2+) channel antagonists can exert opposite effects on Ca(2+) influx, glucose homeostasis and insulin secretion. The influence of the Ca(2+) channel antagonists on pancreatic beta cell functions is dependent on lipophilicity, interactions with the cell membrane lipid bilayer, with SNAREs protein complexes in cell and vesicle membranes, with intracellular receptors, bioavailability and time of elimination from several organs and the bloodstream. In the present work we studied the effect at several doses of new compounds synthesized in the Latvian Institute of Organic Synthesis on blood glucose levels in normal and STZ-induced diabetic rats. The compounds tested were: 1,4-DHP derivatives cerebrocrast (1), etaftoron (2), OSI-1190 (3), OSI-3802 (4), OSI-2954 (5) and known 1,4-DHP derivatives: niludipine (6), nimodipine (7) and nicardipine (8) which possess different lipophilicities. Analysis of the structure-function relationships of the effect of 1,4-DHP derivatives on glucose metabolism showed that cerebrocrast could evoke qualitative differences in activity. Insertion of an OCHF(2) group in position 2 of the 4-phenylsubstituent and propoxyethylgroup R in ester moieties in positions 3 and 5 of the DHP structure, as well as an increase in the number of carbon atoms in the ester moiety, significantly modified the properties of the compound. Thereby cerebrocrast acquired high lipophilicity and membranotropic properties. Cerebrocrast, in a single administration at low doses (0.05 and 0.5 mg x kg(-1), p.o.), significantly decreased the plasma level of glucose in normal rats and in STZ-induced diabetic rats returned plasma glucose to basal levels. This effect was characterized by a slow onset and a powerful long-lasting influence on glucose metabolism, especially in STZ-induced diabetic rats.     

Effect of cerebrocrast on body and organ weights,food and water intake,and urine output of normal rats

Type 2 diabetes is associated with obesity, insulin resistance, hyperglycemia, hyperphagia, polyuria, body weight gain, excessive secretion of glucocorticoids (GCs), thymus involution, adrenal gland hypertrophy, diabetic nephropathy, etc. We examined the effect of cerebrocrast, a new antidiabetic agent (synthesized in the Latvian Institute of Organic Synthesis), on body weight, food and water intake, urine output, and on changes of organ weight: that is, kidney, thymus, adrenal gland of normal rats. Cerebrocrast was administered at doses of 0.05 and 0.5 mg kg⁻¹ per os (p.o.) once a day for three consecutive days, and its effects were observed from 3 to 27 days after the last administration. Cerebrocrast, during the experimental period, decreased body weight by an average of approximately 32.3%, food intake by about 10–15% at the beginning of the experiments and by 22.6% at the end of the experiments, especially at a dose of 0.5 mg kg⁻¹. Water intake and urine output in comparison with controls were decreased. The daily food intake decreased about 1.0 and 2.1 g by administering single cerebrocrast doses of 0.05 and 0.5 mg kg⁻¹ body weight (b.w.), respectively, but by administering for three consecutive days, food intake decreased by about 2.2 and 3.4 g, respectively. The weekly body weight gain decreased by administering a single dose of cerebrocrast by 2.61 and 2.51 g, respectively, and by triple administration it decreased by 4.36 and 3.07 g, respectively. Cerebrocrast has long‐lasting effects on these parameters and on thymus and adrenal gland weight. As cerebrocrast decreased glucose levels in normal and streptozotocin (STZ)‐induced diabetic rats, it also promoted glucose uptake by the brain, intensified insulin action and formation de novo of insulin receptors. We can conclude that cerebrocrast may regulate food intake and body weight through glucose sensing by proopiomelanocortin (POMC) neurons, that are involved in control of glucose homeostasis, stimulation of α‐melanocyte‐stimulating hormone (α‐MSH) secretion, activation of MC4‐Rs and inhibition of neuropeptide Y (NPY) in the ARC of the hypothalamus, affecting the kidney, and causing decreased urine output and water intake. Moreover, it could stimulate secretion of vasopressin. By administration of cerebrocrast thymus mass was increased, thereby preventing the action of GCs. As cerebrocrast inhibited L‐ and T‐type calcium channels, it can prevent vasoconstriction of kidney arterioles and aldosterone secretion that have significant roles in the development of hypertension and diabetic nephropathy. These properties of cerebrocrast are important for treatment of Type 2 diabetes and its consequent development of hypertension and diabetic nephropathy. Copyright © 2008 John Wiley & Sons, Ltd.     

Biological quality metrics: their variability and appropriate scale for assessing streams

The concept of spatial scale is at the research frontier in ecology, and although focus has been placed on trying to determine the role of spatial scale in structuring communities, there still is a further need to standardize which organism groups are to be used at which scale and under which circumstances in environmental assessment. This paper contributes to the understanding of the variability at different spatial scales (reach, stream, river basin) of metrics characterizing communities of different biological quality elements (macrophytes, fishes, macroinvertebrates and benthic diatoms) as defined by the Water Framework Directive. For this purpose, high-quality reaches from medium-sized lowland streams of Latvia, Ecoregion 15 (Baltic) were sampled using a nested hierarchical sampling design: (river basin → stream → reach). The variability of metrics within the different groups of biological quality elements confirmed that large-bodied organisms (macrophytes and fish) were less variable than small-bodied organisms (macroinvertebrates and benthic diatoms) at reach, stream and river basin scales. Single metrics of biological quality elements had the largest variation at the reach scale compared with stream and basin scales. There were no significant correlations between biodiversity indices of the different organism groups. The correlation between diversity indices (Shannon’s and Simpson’s) of the biological quality elements (macrophytes, fish, benthic macroinvertebrates and benthic diatoms) and a number of measured environmental variables varied among the different organism groups. Relationships between diversity indices and environmental factors were established for all groups of biological quality elements. Our results showed that metrics of macrophytes and fish could be used for assessing ecological quality at the river basin scale, whereas metrics of macroinvertebrates and benthic diatoms were most appropriate at a smaller scale.     

Acute effect of antidiabetic 1,4-dihydropyridine compound cerebrocrast on cardiac function and glucose metabolism in the isolated, perfused normal rat heart

Diabetes mellitus (DM) is an important cardiovascular risk factor and is associated with abnormalities in endothelial and vascular smooth muscle cell function, evoked by chronic hyperglycemia and hyperlipidemia. Chronic insulin deficiency or resistance is marked by decreases in the intensity of glucose transport, glucose phosphorylation, and glucose oxidation, plus decreases in ATP levels in cardiac myocytes. It is important to search for new agents that promote glucose consumption in the heart and partially inhibit extensive fatty acid beta-oxidation observed in diabetic, ischemia. When the oxygen supply for myocardium is decreased, the heart accumulates potentially toxic intermediates of fatty acid beta-oxidation, that is, long-chain acylcarnitine and long-chain acyl-CoA metabolites. Exogenous glucose and heart glycogen become an important compensatory source of energy. Therefore we studied the effect of the antidiabetic 1,4-dihydropyridine compound cerebrocrast at concentrations from 10(-10) M to 10(-7) M on isolated rat hearts using the method of Langendorff, on physiological parameters and energy metabolism. Cerebrocrast at concentrations from 10(-10) M to 10(-7) M has a negative inotropic effect on the rat heart. It inhibits L-type Ca(2+)channels thereby diminishing the cellular Ca(2+) supply, reducing contractile activity, and oxygen consumption, that normally favors enhanced glucose uptake, metabolism, and production of high-energy phosphates (ATP content) in myocardium. Cerebrocrast decreases heart rate and left ventricular (LV) systolic pressure; at concentrations of 10(-10) M and 10(-9) M it evokes short-term vasodilatation of coronary arteries. Increase of ATP content in the myocytes induced by cerebrocrast has a ubiquitous role. It can preserve the integrity of the cell plasma membranes, maintain normal cellular function, and inhibit release of lactate dehydrogenase (LDH) from cells that is associated with diabetes and heart ischemia. Administration of cerebrocrast together with insulin shows that both compounds only slightly enhance glucose uptake in myocardium, but significantly normalize the rate of contraction and relaxation ( +/- dp/dt). The effect of insulin on coronary flow is more pronounced by administration of insulin together with cerebrocrast at a concentration of 10(-7) M. Cerebrocrast may promote a shift of glucose consumption from aerobic to anerobic conditions (through the negative inotropic properties), and may be very significant in prevention of cardiac ischemic episodes.     

Effect of cerebrocrast on the function of human platelets and release of the arachidonic acid from plasma membrane

Diabetes mellitus (DM) is accompanied by several cardiovascular complications such as coronary artery disease, atherosclerosis, hypertension, cerebral and myocardial infarction, etc. DM induces the alteration of platelet functions including activation, hyperaggregation, adhesiveness, and formation of thrombi. Release of AA from phospholipids of the PM, synthesis of TxA(2),PGE(2), activity of PLA(2), and PLC are increased in the platelets of the DM patients. Stimulation of PLA(2) activity and accumulation of bioactive metabolites such as AA, its oxygenated derivatives, prostaglandins and PAF can evoke glucose production, also. In this study we explored the effect of the 1,4-dihydropyridine compound cerebrocrast at a low concentration (10(-6)-10(-8)M) on the level of intracellular calcium in unstimulated human platelets and those stimulated with thrombin as well as release of [(3)H] AA from phospholipids of platelet PM. Cerebrocrast at a concentration of 10(-6) M decreased the basal level of intracellular calcium concentration (platelets were loaded with Fura-2) in unstimulated as well as in thrombin stimulated platelets. Cerebrocrast at concentrations of 10(-6), 10(-7), 10(-8) M inhibited release of [(3)H] AA from phospholipids of platelet PM. We conclude that blockade of human platelet activation with cerebrocrast can prevent aggregation, adhesion and formation of thrombi. The inhibition of [(3)H] AA release from phospholipids of platelet PM can prevent formation of eicosanoids such as TxA(2), PGG(2), and PGH(2) plus AA oxygenated derivatives. These effects of cerebrocrast are very significant in the treatment of DM-evoked cardiovascular complications.     

Effect of cerebrocrast, a new long-acting compound on blood glucose and insulin levels in rats when administered before and after STZ-induced diabetes mellitus

Insulin-dependent diabetes mellitus (IDDM) is an autoimmune disease that is characterized by selective destruction of insulin secreting pancreatic islets beta-cells. The formation of cytokines (IL-1beta, IL-6, TNF-alpha, etc.) leads to extensive morphological damage of beta-cells, DNA fragmentation, decrease of glucose oxidation, impaired glucose-insulin secretion and decreased insulin action and proinsulin biosynthesis. We examined the protective effect of a 1,4-dihydropyridine (DHP) derivative cerebrocrast (synthesized in the Latvian Institute of Organic Synthesis) on pancreatic beta-cells in rats possessing diabetes induced with the autoimmunogenic compound streptozotocin (STZ). Cerebrocrast administration at doses of 0.05 and 0.5 mg/kg body weight (p.o.) 1 h or 3 days prior to STZ as well as at 24 and 48 h after STZ administration partially prevented pancreatic beta-cells from the toxic effects of STZ, and delayed the development of hyperglycaemia. Administration of cerebrocrast starting 48 h after STZ-induced diabetes in rats for 3 consecutive days at doses of 0.05 and 0.5 mg/kg body weight (p.o.) significantly decreased blood glucose level, and the effect remained 10 days after the last administration. Moreover, in these rats, cerebrocrast evoked an increase of serum immunoreactive insulin (IRI) level during 7 diabetic days as compared to both the control normal rats and the STZ-induced diabetic control rats. The STZ-induced diabetic rats that received cerebrocrast had a significantly high serum IRI level from the 14th to 21st diabetic days in comparison with the STZ-induced diabetic control.The IRI level in serum as well as the glucose disposal rate were significantly increased after stimulation of pancreatic beta-cells with glucose in normal rats that received cerebrocrast, administered 60 min before glucose. Glucose disposal rate in STZ-induced diabetic rats as a result of cerebrocrast administration was also increased in comparison with STZ-diabetic control rats. Administration of cerebrocrast in combination with insulin intensified the effect of insulin. The hypoglycaemic effect of cerebrocrast primarily can be explained by its immunomodulative properties. Moreover, cerebrocrast can act through extrapancreatic mechanisms that favour the expression of glucose transporters, de novo insulin receptors formation in several cell membranes as well as glucose uptake.