Effects of dexamethasone and chlorpromazine treatment on X-Y dissociation and multinucleated giant cell formation in hyperthermic mice

The effects of anti-stress drugs on X-Y dissociation and multinucleated giant cell formation in the testes of hyperthermic mice were determined for the possible use of such drugs in animal production. Mice were injected with dexamethasone 1 mg/kg or 2 mg/kg, chlorpromazine 1 mg/kg or 4 mg/kg, or 0.85% saline, and heat stressed for 4 d at 35 +/- 1 C and 65 +/- 1 % relative humidity. Mice were killed 5 d after stress, and the testes processed for observation of meiotic chromosomes and testicular histology. Heat stress caused a significant increase in X-Y bivalent dissociation in diakinesis-metaphase I spermatocytes and a significant increase in formation of multinucleated giant cells. Drug treatment increased X-Y dissociation but had no significant effect on giant cell formation.     

Pharmacologic protection of perfused rat heart against global ischemia

Using a Langendorff rat heart model, studies were performed on the effects of three drugs in protecting the heart against global ischemia. The drugs used were: (a) MR-256, a prostaglandin oligomeric derivative, which is a calcium chelating agent and at the same time, is an inhibitor of phospholipase A2 activity, (b) chlorpromazine which is not a calcium chelator, but is a calmodulin antagonist and is an inhibitor of phospholipase A2 activity, and (c) BAPTA/AM, a calcium chelating agent, but which is not an inhibitor of phospholipase A2 activity. The perfused heart was exposed to 15 minutes of global ischemia. In control experiments (no drug), the ventricular pressure recovered to 26.4 +/- 6.7% (n = 22) of the original level. With pretreatment of (a) MR-256 (b) chlorpromazine, and (c) BAPTA/AM, maximum recoveries were 0.5 +/- 6.7% (n = 5), 88.7 +/- 8.5% (n = 5), 45.3 +/- 26.6% (n = 5), respectively. MR-256 and chlorpromazine were found to react with free radicals. The modes of action of these three different types of drugs are discussed.     

The effect of calmodulin and chlorpromazine on the activity of phospholipases A2 from various sources

The effects of calmodulin and chlorpromazine on purified phospholipase A2 preparations from snake venoms: cobra (Naja naja oxiana), echis (Ehis multisquamatus) and Agkistrodon halys halys, as well as on phospholipases A2 from rat liver mitochondria and human platelets were studied. It was shown that within the concentration range of 1-5 microM calmodulin stimulates the phospholipase activity. Chlorpromazine inhibits the activity of these enzymes, the degree of inhibition being different for various phospholipases. Calmodulin was shown to interact with the phospholipases in the absence of exogenous Ca2+. The results obtained indicate that all phospholipases tested are calmodulin-dependent enzymes.     

Vanadium and cadmium in vivo effects in teleost cardiac muscle: metal accumulation and oxidative stress markers

Several biological studies associate vanadium and cadmium with the production of reactive oxygen species (ROS), leading to lipid peroxidation and antioxidant enzymes alterations. The present study aims to analyse and compare the oxidative stress responses induced by an acute intravenous exposure (1 and 7 days) to a sub-lethal concentration (5 mM) of two vanadium solutions, containing different vanadate n-oligomers (n=1-5 or n=10), and a cadmium solution on the cardiac muscle of the marine teleost Halobatrachus didactylus (Lusitanian toadfish). It was observed that vanadium is mainly accumulated in mitochondria (1.33+/-0.26 microM), primarily when this element was administrated as decameric vanadate, than when administrated as metavanadate (432+/-294 nM), while the highest content of cadmium was found in cytosol (365+/-231 nM). Indeed, decavanadate solution promotes stronger increases in mitochondrial antioxidant enzymes activities (catalase: +120%; superoxide dismutase: +140%) than metavanadate solution. On contrary, cadmium increases cytosolic catalase (+111%) and glutathione peroxidases (+50%) activities. It is also observed that vanadate oligomers induce in vitro prooxidant effects in toadfish heart, with stronger effects induced by metavanadate solution. In summary, vanadate and cadmium are differently accumulated in blood and cardiac subcellular fractions and induced different responses in enzymatic antioxidant defence mechanisms. In the present study, it is described for the first time the effects of equal doses of two different metals intravenously injected in the same fish species and upon the same exposure period allowing to understand the mechanisms of vanadate and cadmium toxicity in fish cardiac muscle.     

Effects of diallyl sulfide and zinc on testicular steroidogenesis in cadmium-treated male rats

Cadmium (Cd) is one of the environmental pollutants that affect various tissues and organs including testis. Harmful effect of cadmium on testis is known to be germ cell degeneration and impairment of testicular steroidogenesis. In the present study, the effect of diallyl sulfide (DAS), a sulfur-containing volatile compound present in garlic, and zinc (Zn) was investigated on cadmium-induced testicular toxicity in rats. Male adult Wistar rats treated with cadmium (2.5 mg/kg body wt, five times a week for 4 weeks) showed decreased body weight, paired testicular weight, relative testicular weight, serum testosterone, luteinizing hormone, follicle-stimulating hormone, and testicular total antioxidant capacity (TAC) and protein levels. Testicular steroidogenic enzymes, such as 3beta-hydroxysteroid dehydrogenase (3beta-HSD) and 17beta-hydroxysteroid dehydrogenase (17beta-HSD), and marker enzymes, such as sorbitol dehydrogenase (SDH), lactate dehydrogenase (LDH), acid phosphatase (ACP), alkaline phosphatase (ALP), and glucose-6-phosphate dehydrogenase (G6PD), showed a significant decrease in activities whereas that of gamma-glutamyl transferase was significantly increased after cadmium exposure. The results have revealed that concurrent treatment with DAS or zinc restored key steroidogenic enzymes, SDH, LDH, and G6PD and increased testicular weight significantly. DAS restored the TAC level and increased testosterone level and relative testicular weight significantly. Zinc restored testicular protein level and body weight. It can be concluded that cadmium causes testicular toxicity and inhibits androgen production in adult male rats probably by affecting pituitary gonadotrophins and that concurrent administration of DAS or zinc provides protection against cadmium-induced testicular toxicity.     

Involvement of calcium signaling in the fibronectin-stimulated macrophage recognition of oxidatively damaged erythrocytes

Macrophages recognize oxidatively damaged autologous erythrocytes, and cell surface fibronectin of macrophages enhances the recognition (Beppu et al., FEBS Lett. 295 (1991) 135-140). In the present study, mechanisms of enhanced macrophage recognition of oxidatively damaged erythrocytes by fibronectin were investigated. Monolayers of thioglycollate-induced mouse peritoneal macrophages with cell surface fibronectin recognized autologous erythrocytes oxidized with an iron catalyst ADP/Fe(3+). The macrophage recognition of the oxidized erythrocytes was inhibited partially by pretreatment of the macrophage monolayers with a Ca(2+) channel blocker (diltiazem), calmodulin inhibitors (W-7, trifluoperazine, chlorpromazine and dibucaine), an inhibitor of myosin light chain kinase (ML-9), a microfilament formation inhibitor (cytochalasin B), phospholipase A(2) inhibitors (4-bromophenacyl bromide, mepacrine) and cyclooxygenase inhibitors (indomethacin and aspirin). Monolayers of macrophages depleted of fibronectin by trypsinization lost the ability of recognizing oxidized erythrocytes, but acquired the ability when stimulated with a fibronectin-coated coverslip. The recognition of fibronectin-stimulated trypsinized macrophages was also inhibited by the above inhibitors. On treatment with Ca ionophore A23187, trypsinized macrophages acquired the ability to recognize oxidized erythrocytes. The recognition of Ca ionophore-stimulated trypsinized macrophages was inhibited by the above inhibitors except the Ca(2+) channel blocker. These results indicate that the Ca(2+) signaling including Ca(2+) influx, calmodulin activation and myosin light chain phosphorylation are involved in the fibronectin stimulation of the recognition of macrophages for oxidized erythrocytes. Involvement of microfilament formation and arachidonate cascade in the fibronectin stimulation was also suggested.     

Cadmium induced limb defects in mice: strain associated differences in sensitivity.

Cadmium (CdSO4) was given ip on day 9 at 12 or 24 mumol/kg to pregnant CD-1 (non-inbred) mice. Fetuses showed malformations of the limbs, face, trunk, and tail. There was a statistically significant relationship between the dose of cadmium and the malformation rate. Cadmium (12 mumol/kg ip on day 9) was then given to mice of six inbred strains three of which (A/J, BALB/cJ, and C57BL6J) carry a gene cdm for resistance to cadmium-induced testicular damage, and three strains (AKR/J, CBA/J, and DBA/2J) which do not. Paradoxically, the three strains resistant to cadmium induced testicular damage were significantly more sensitive to its teratogenic effects than were the other three strains. In all inbred strains most malformations involved the limbs. All forelimb defects found in inbred or non-inbred cadmium treated mice were postaxial and indistinguishable from those produced by acetazolamide in mice. The remarkable similarity of the cadmium- and acetazolamide-induced forelimb malformations may be a reflection of the limited number of ways that a rodent forelimb can react to a teratogenic insult. The hindlimb defects were all preaxial.     

Effects of calcium, calmodulin, protein kinase C and protein tyrosine kinases on volume-activated taurine efflux in human erythroleukemia cells.

The effects of calcium, calmodulin, protein kinase C (PKC) and protein tyrosine kinase (PTK) modulators were examined on the volume-activated taurine efflux in the erythroleukemia cell line K562. Exposure to hypoosmotic solution significantly increased taurine efflux and intracellular calcium concentration ([Ca2+]i). The Ca2+ channel blockers La3+ (1 mM), verapamil (200 microM) and nifedipine (100 microM) inhibited the hypoosmotically-induced [Ca2+]i increase by more than 90%, while the volume-activated taurine efflux was inhibited by 61.3 +/- 9.5, 74.1 +/- 9.3 and 38.0 +/- 1.5%, respectively. Furthermore, the calmodulin inhibitors W7 (50 microM) and trifluoperazine (10 microM) and the Ca2+/calmodulin-dependent protein kinase II inhibitor KN-62 (2 microM) significantly blocked the volume-activated taurine efflux by 93.4 +/- 2.7, 77.9 +/- 3.5 and 61.3 +/- 15.8%, respectively. In contrast, the PKC inhibitor staurosporine (200 nM) or the PKC activator phorbol 12-myristate 13-acetate (100 nM) did not have significant effects on the volume-activated taurine efflux. However, pretreatment with PTK inhibitors genistein, tyrphostin A25, and tyrphostin A47 blocked the volume-activated taurine efflux. These results suggest that the volume-activated taurine efflux in K562 cells may not directly involve Ca2+, but may require the presence of calmodulin and/or PTK.     

Effects of ketoconazole on rat testicular steroidogenic enzymatic activities

Ketoconazole (K) is an antifungal imidazole derivative which has been shown to be a potent inhibitor of testosterone (T) biosynthesis in rodents and humans. To study the effect of K on rat testicular steroidogenesis we measured the activities of five testicular microsomal steroidogenic enzymes in K-treated rats and controls. Thirty male adult rats were given either 2 mg K or water every 12 hours by mouth during 5 days. Mean testicular weight was similar in both groups of animals. The K-treated group had a T serum concentration of 83 +/- 14 ng/dL whereas it was 94 +/- 16 ng/dL in the control group (NS). The K-treated animals had decreased activities of the 3 beta-hydroxysteroid dehydrogenase (830 +/- 48 vs 2,245 +/- 109 pmol/mg protein/min, P less than 0.001), 17-hydroxylase (243 +/- 5 vs 676 +/- 17 pmol/mg protein/min, P less than 0.001), 17-ketosteroid reductase (31 +/- 2 vs 169 +/- 7 pmol/mg protein/min, P less than 0.001), and aromatase enzymes (92 +/- 6 vs 123 +/- 7 pmol/mg protein/min, P less than 0.01). The 17,20-desmolase activity was similar in both groups of animals (210 +/- 4 vs 171 +/- 18 pmol/mg protein/min). We conclude that K given orally to rats inhibits the activity of several testicular steroidogenic enzymes.     

Modulatory effects of lipoic acid and selenium against cadmium-induced biochemical alterations in testicular steroidogenesis

Exposure to toxic metals including cadmium has become an increasingly recognized source of illness worldwide. Cadmium (Cd(2+) ) is one of the environmental pollutants affecting various tissues and organs including testis. The protective effect of lipoic acid and selenium on Cd(2+) -induced testicular damage was investigated. Accordingly, male Wistar rats were allocated into four groups (n = 8; each). Gp I: (control), whereas the other 3 groups received CdCl(2) (2 mg/kg, i.p. for 28 days) alone or in combination with either (i) lipoic acid (35 mg/kg, p.o) or (ii) selenium (0.35 mg/kg, p.o) throughout the experiment. Serum testosterone, luteinizing hormone and follicle-stimulating hormone levels significantly decreased in the Cd(2+) -exposed rats. The activities of testicular key androgenic enzymes, 3β-hydroxysteroid dehydrogenase and 17 β-HSD significantly decreased in Cd(2) exposed rats compared to the control counterparts. In addition, the activities of testicular marker enzymes were significantly altered in cadmium-treated animals. Significant reductions in body and testicular weight as well as antioxidant status were also observed in Cd(2+) -exposed rats. Moreover, some testicular metal levels were altered. Lipoic acid and selenium significantly increased serum testosterone level and restored testicular activity of 3β-HSD and 17 β-HSD and were effective in modulation of most of the measured biochemical parameters. The biochemical parameters were further confirmed with histopathological findings. In conclusion, the present study demonstrated the beneficial influences of lipoic acid and selenium in reducing harmful effects of Cd(2+) in rats' testes.     

Inhibition of growth and of thymidine incorporation into DNA in Tetrahymena by chlorpromazine, pimozide and penfluridol

The antipsychotic drugs chlorpromazine, pimozide, and penfluridol caused a 50% inhibition of growth of Tetrahymena at concentrations of 4.5, 5.5, and 1.5 microM, respectively. The degree of growth inhibition was dependent on the concentration of cells; higher drug concentrations were needed to produce inhibition of denser cell cultures. Binding studies with penfluridol showed that 50% growth inhibition resulted when approximately 50 mumoles of drug were bound per 10(6) cells. A 20-min preincubation of cells with chlorpromazine (14.7 microM) inhibited DNA synthesis by 46%, and with penfluridol (4 microM) DNA synthesis was inhibited by 27%. The incorporation of labeled thymidine into the thymidine triphosphate pool was inhibited by chlorpromazine but not by penfluridol, indicating that the drugs produce their growth inhibitory effects by different mechanisms. TDP kinase activity was demonstrated in a particle-free fraction of the cells. Its enzymatic activity was not affected by added chlorpromazine, penfluridol, or calmodulin, suggesting that inhibition of DNA synthesis by these drugs may be a consequence of growth inhibition.     

Resistance of KP and CBA mice to cadmium chloride assessed on the basis of the bone marrow stromal cell growth in vitro

Using a culture of bone marrow stromal cells, resistance of two inbred mice strains, KP and CBA, to cadmium chloride was assessed. Mice were administered with a single dose of CdCl2 (12 mumoles (kg b. wt) and bone marrow cells were examined after 7 days of culture. In KP strain, a decrease in the number of fibroblasts and proliferation of adipocytes was observed, as compared with control animals. In CBA strain, cadmium did not influence the number of fibroblasts and caused only an insignificant increase in the amount of adipocytes. Red blood cell count, hematocrit and Fe2+ level were unchanged both strains after cadmium administration. The observed differences in populations of cultured stromal cells confirmed the sensitivity of the KP mice to cadmium and indicated that the stromal cells can be regarded as useful indicator of cadmium intoxication.     

Effects of calmodulin and lipoxygenase inhibitors on LH (lutropin)- and LHRH (luliberin)-agonist-stimulated steroidogenesis in rat Leydig cells.

The results of this study, carried out with purified rat Leydig cells, indicate that there are no major differences in the stimulating effects of lutropin (LH) and luliberin (LHRH) agonists on steroidogenesis via mechanisms that are dependent on Ca2+. This was demonstrated by using inhibitors of calmodulin and the lipoxygenase pathways of arachidonic acid metabolism. All three calmodulin inhibitors used (calmidazolium, trifluoperazine and chlorpromazine) were shown to block LH- and LHRH-agonist-stimulated steroidogenesis. This probably occurred at the step of cholesterol transport to the mitochondria. Similarly, three lipoxygenase inhibitors (nordihydroguaiaretic acid, BW755c and benoxaprofen), inhibited both LH- and LHRH-agonist-stimulated steroidogenesis. The amounts of the inhibitors required were similar for LH- and LHRH-agonist-stimulated steroidogenesis. Steroidogenesis stimulated by the Ca2+ ionophore A23187 was also inhibited, but higher concentrations of the inhibitors were required. Indomethacin (a cyclo-oxygenase inhibitor) increased LHRH-agonist-stimulated steroidogenesis;this is consistent with the role of the products of arachidonic acid metabolism via the alternative, lipoxygenase, pathway. The potentiation of LH-stimulated testosterone production by LHRH agonist was unaffected by indomethacin or by lipoxygenase inhibitors at concentrations that inhibited LH-stimulated testosterone production by 75-100%. It was not possible to eliminate a role of calmodulin in modulating the potentiation, although higher concentrations of the inhibitors were generally required to negate the potentiation than to inhibit LH- or LHRH-agonist-stimulated testosterone production.     

Effects of calcium ionophore, A23187 and calmodulin inhibitors on intercellular communication in the rat myometrium

We have studied the effect of a calcium ionophore, A23187, and the purported calmodulin inhibitors, calmidazolium and chlorpromazine, on direct intercellular communication between smooth muscle cells in the myometrium of delivering rats. The extent of cell-to-cell coupling was determined by exposing one portion of small strips of longitudinal myometrium to 2-[3H] deoxy-D-glucose (2-DG) and determining the distribution and apparent diffusion coefficient (Da) for this tracer after a 5-h period for diffusion. The distribution and Da for 2-DG were significantly (p less than 0.05) reduced by exposure to A23187 in Krebs-Ringer solution with 2.5 mM Ca++, partially reduced in Krebs solution with A23187 and low Ca++ (1-10 microM), but the drug had no effect when used with Ca++-free solutions with [ethylenebis (oxyethylene-nitrilo)] tetraacetic acid (EGTA). The calmodulin inhibitors blocked the effects of A23187 in a dose-dependent fashion, and at higher concentrations, the extent of 2-DG diffusion was not different from that in control tissues. Surprisingly, however, a dose-dependent reduction in coupling was also observed in tissues exposed to the calmodulin inhibitors alone. Structural studies failed to reveal any change in the area of gap junctions between the myometrial cells following the above treatments, suggesting that the reduced exchange of 2-DG resulted from a decrease in the permeability of gap junctions between the muscle fibers.     

A possible regulatory role of Ca++-calmodulin system in cellular cholesterol ester hydrolysis in the steroidogenic response to ACTH in bovine adrenocortical cells

In order to corroborate the regulatory role of Ca++-calmodulin system in the steroidogenic response to adrenocorticotropic hormone (ACTH), the effects of calmodulin inhibitors (chlorpromazine, trifluoperazine, and W-7) on cortisol production and cellular cholesterol ester hydrolysis induced by ACTH in bovine adrenocortical cells were examined. Three calmodulin inhibitors diminished not only the cholesterol ester hydrolysis and cortisol production induced by ACTH in the presence of Ca++, but also inhibited the Ca++-induced hydrolysis and cortisol production in the absence of ACTH. Neither cortisol production in crude mitochondrial fraction nor the ACTH-induced Ca++-influx was affected by chlorpromazine. These results indicate that Ca++f-calmodulin system plays a significant regulatory role in the supply of free cholesterol to the adrenal mitochondria in the steroidogenic response to ACTH.     

Spermatozoa output, testicular sperm reserve and epididymal storage capacity of the Red Sokoto goats indigenous to northern Nigeria

Fifteen Red Sokoto goats (bucks) aged 2 to 2 1 2 years with a mean body weight of 17.80+/-1.24 kg were used for studying daily spermatozea output, testicular and epididymal sperm reserves. Semen ejaculates were obtained from each buck and evaluated daily for six weeks. Semen characteristics were as follows: volume, 0.72+/-0.91ml; colour, milky/creamy; sperm concentration/ml, 0.61+/-0.05 x 10(9); mass motility, 3.80+/-0.33; individual metility, 77.50+/-5.60; and normal sperm morphology, 80.00+/-10.50. Mean scretal circumference was 21.80+/-0.29. Mean testicular and spididymal (capita, corpora and caudas) weights (gm) were 83.74+/-5.33, 7.27+/-0.63, 2.22+/-0.23 and 6.13+/-0.73 respectively. Corresponding sperm reserves were 44.32+/-5.79, 8.82+/-1.95, 4.99+/-0.86 and 45.64+/-7.87 x 10(9). Scrotal circumference was positively correlated to testicular weight (r = 0.86), testicular spermatids (r = 0.77), and caudas spermatozoa (r = 0.83).     

The nitric oxide prodrug, V-PYRRO/NO, protects against cadmium toxicity and apoptosis at the cellular level.

The liver is an important target tissue of cadmium. The compound O2-vinyl 1-(pyrrolidin-1-yl)diazen-1-ium-1,2 diolate (V-PYRRO/NO) is a liver-selective nitric oxide (NO) prodrug that is metabolized by hepatic P450 enzymes to release NO in hepatocytes. In vivo, V-PYRRO/NO can protect against the toxicity of various hepatotoxicants, including cadmium. Since NO is an effective vasodilator, whether this protective effect against cadmium toxicity is at the level of the hepatic vascular system or actually within the liver cells has not been defined. Thus, we studied the effects of V-PYRRO/NO pretreatment on cadmium-induced toxicity and apoptosis in cultured rat liver epithelial (TRL 1215) cells. Cells were pretreated with V-PYRRO/NO at 500 or 1000 microM for up to 24 h, then exposed to cadmium (as CdCl2) for additional 24 h and cytotoxicity was measured. Cadmium was significantly less cytotoxic in V-PYRRO/NO (1000 microM) pretreated cells (LC50=6.1+/-0.6 microM) compared to control cells (LC50=3.5+/-0.4 microM). TRL 1215 cells acted upon the prodrug to release NO, producing nitrite levels in the extracellular media after 24 h of exposure to 500 or 1000 microM V-PYRRO/NO measured at 87.0+/-4.2 and 324+/-14.8 microM, respectively, compared to basal levels of 7.70+/-0.46 microM. V-PYRRO/NO alone produced small increases in metallothionein (MT), a metal-binding protein associated with cadmium tolerance. However, V-PYRRO/NO pretreatment greatly enhanced cadmium induction of MT. V-PYRRO/NO pretreatment also markedly reduced apoptotic cell death induced by cadmium (5 microM), apparently by blocking cadmium-induced activation of the c-Jun N-terminal kinase (JNK) pathway. Thus, the prodrug, V-PYRRO/NO, protects against the adverse effects of cadmium directly within rat liver cells apparently through generation of NO and, at least in part, by facilitation of cadmium-induced MT synthesis.     

Probiotics as a Complementary Therapy in the Model of Cadmium Chloride Toxicity: Crosstalk of β-Catenin,BDNF, and StAR Signaling Pathways

Cadmium chloride (CdCl₂) is a ubiquitous environmental toxicant that causes a variety of disturbances in biological systems, including brain dysfunction and testicular tissue degeneration. On the other hand, it is supposed that beneficial properties of probiotic bacteria (Lactobacillus and Acidobacillus) are related to their capacity to adhere or bind different targets, thus leading to improved intestinal microbial balance and other benefits to the host. Bearing aforementioned in mind, the present study was undertaken to investigate the protective effect of probiotic supplementation against cadmium chloride-induced brain and testis toxicity in mice model. Animals received Lactobacillus and Acidobacillus either alone or added to folic acid for 1 week before CdCl2 intoxication in a dose of 20 mg/kg BW followed by probiotics (5?×?10⁹⁾ and/or folic acid (12 mg/kg) treatment for 3 weeks. The levels of malondialdehyde (MDA), butyrl choline esterase (BCHE), reduced glutathione (GSH), and total superoxide dismutase (SOD) activities were investigated. Finally, cadmium neurotoxicity was determined by estimating the gene expression of β-catenin and brain-derived neurotrophic factor (BDNF), as well as estimating the alterations in testicular function by determining acid phosphatase level in addition to steroidogenic acute regulatory protein (StAR) and 17-hydroxy steroid dehydrogenase (17-β HSD) gene expression. Based on our results, we can conclude that exposure of mice to cadmium chloride resulted in a significant elevation in MDA, BCHE levels accompanied with a significant reduction in GSH and SOD activities compared to the control value. CdCl₂ also downregulated the gene expression of β-catenin and BDNF, as well as acid phosphatase level, in addition to StAR and 17-β HSD gene expression. These deviated parameters were significantly modulated in the co-treated animals with probiotics compared with the cadmium-treated group. In conclusion, Lactobacillus and folic acid in a mixture with cadmium acted beneficially to an organism, increasing the cadmium excretion in feces, and consequently increasing β-catenin and BDNF in brain tissue and StAR and 17-β HSD in testis and improving their functions. Histoarchitecture analysis confirmed these results.     

Characterization of the autophosphorylation of chicken gizzard caldesmon.

Caldesmon, an actin- and calmodulin-binding protein of smooth muscle, is a protein serine/threonine kinase capable of Ca2+/calmodulin-dependent autophosphorylation [Scott-Woo & Walsh (1988) Biochem. J. 252, 463-472]. Phosphorylation nullifies the inhibitory effect of caldesmon on the actin-activated Mg2+-ATPase activity of smooth-muscle myosin [Ngai & Walsh (1987) Biochem. J. 244, 417-425]. We have characterized the kinase activity of caldesmon of chicken gizzard smooth muscle. Autophosphorylation requires Ca2+/calmodulin, but is unaffected by other second messengers (Ca2+/phospholipid/diacylglycerol, cyclic AMP or cyclic GMP), and is inhibited by the calmodulin antagonists chlorpromazine and compound 48/80, with 50% inhibition at 39.8 microM and 12.0 ng/ml respectively. Half-maximal activation of autophosphorylation occurs at 60-80 nM-Ca2+ and 0.14 microM-calmodulin, and maximal activity at 0.14-0.18 microM-Ca2+ and 1 microM-calmodulin; activation is gradually lost at higher Ca2+ and calmodulin concentrations. Autophosphorylation is pH-dependent, with maximal activity over the range pH 7-9, and requires free Mg2+ in addition to the MgATP2- substrate. The Km for ATP is 15.6 +/- 4.1 microM (mean +/- S.D., n = 4), and kinase activity is inhibited by increasing ionic strength [half-maximal inhibition at I = 0.094 +/- 0.009 M (mean +/- S.D., n = 4)]. Autophosphorylation does not affect the rate of hydrolysis of caldesmon (free or bound to calmodulin) by alpha-chymotrypsin. However, a slight difference in peptides generated from phospho- and dephospho-forms of caldesmon is observed. The binding of phospho- or dephospho-caldesmon to F-actin protects the protein against chymotryptic digestion, but does not alter the pattern of peptide generation. Characterization of proteolytic fragments of caldesmon generated by alpha-chymotrypsin and Staphylococcus aureus V8 protease enables localization of the phosphorylation sites and the kinase active site within the caldesmon molecule.