Zinc deficiency decreases the activity of calmodulin regulated cyclic nucleotide phosphodiesterases in vivo in selected rat tissues

The effect of zinc deficiency on calmodulin function was investigated by assessing the in vivo activity of two calmodulin regulated enzymes, adenosine 3′,5′-monophosphate (c-AMP) and guanosine 3′,5′-monophosphate (c-GMP) phosphodiesterase (PDE) in several rat tissues. Enzymatic activities in brain, heart, and testis of rats fed a zinc deficient diet were compared with activities in these tissues from pair fed, zinc supplemented rats. In testis, a tissue in which zinc concentration decreased with zinc deficient diet, enzyme activities were significantly decreased over those in rats who were pair fed zinc supplemented diets. In brain and heart, tissues in which zinc concentrations did not change with either diet, enzymatic activities between the groups were not different. These results indicate that zinc deficiency influences the activity of calmodulin-regulated phosphodiesterases in vivo supporting the hypothesis that zinc plays a role in calmodulin function in vivo in zinc sensitive tissues.     

In vivo effects of cadmium on calmodulin and calmodulin regulated enzymes in rat brain.

Effect of chronic cadmium (Cd) exposure and the influence of diethyldithiocarbamate (DDC) on Cd absorption was studied on the brain of young male Wistar rats. A significant amount of Cd accumulated in cerebral cortices of rats after 4 weeks of Cd (6 mg/kg body wt) exposure (through gastric intubation). The biological activity of calmodulin (CaM) decreased significantly (p less than 0.001) in the cerebral cortices of these animals in comparison to the control group. 3'-5' Phosphodiesterase and synaptic membrane Ca(2+)-Mg(2+) ATPase were also significantly affected (p less than 0.01 and p less than 0.001 respectively). However, Cd treatment did not alter synaptic membrane adenylate cyclase activity and DDC (9.2 mg/kg body wt, intraperitoneal) treatment along with Cd (6 mg/kg body wt) enhanced Cd accumulation in cerebral cortices of treated animals resulting in an increased inhibition of CaM and CaM dependent enzymes. These data suggest that Cd may be acting via binding to CaM and uncoupling it from its normal cellular control of calcium.     

In vitro effects of toxaphene on mitochondrial calcium ATPase and calcium uptake in selected rat tissues

$\text{In vitro}$ effects of toxaphene on Ca²⁺-ATPase activity and ⁴⁵Ca²⁺-uptake were studied in mitochondrial fractions of heart, kidney and liver tissues of rat. Mitochondrial fractions were prepared by the conventional centrifugation method. Ca²⁺-ATPase activity was determined by measuring the inorganic phosphate liberated during ATP hydrolysis. Toxaphene inhibited Ca²⁺-ATPase in a concentration dependent manner in all the three tissues. Substrate activation kinetics, with heart, kidney and liver tissue fractions, revealed that toxaphene inhibited Ca²⁺-ATPase activity non-competetively by decreasing the maximum velocity of the enzyme without affecting the enzyme-substrate affinity. Toxaphene also inhibited mitochondrial ⁴⁵Ca²⁺-uptake in the three selected tissues in a concentration dependent manner. These results indicate that toxaphene is an inhibitor of mitochondrial Ca²⁺-ATPase and calcium transport in heart, kidney and liver tissues of rat.     

The response of zinc transporter gene expression of selected tissues in a pig model of subclinical zinc deficiency

This study compared the relative mRNA expression of all mammal zinc (Zn) transporter genes in selected tissues of weaned piglets challenged with short-term subclinical Zn deficiency (SZD). The dietary model involved restrictive feeding (450 g/animal*day⁻¹) of a high-phytate diet (9 g/kg) supplemented with varying amounts of zinc from ZnSO₄*7H₂O ranging from deficient to sufficient supply levels (total diet Zn: 28.1, 33.6, 38.8, 42.7, 47.5, 58.2, 67.8, 88.0 mg Zn/kg). Total RNA preparations comprised jejunal and colonic mucosa as well as hepatic and nephric tissue. Statistical modelling involved broken-line regression (P≤.05). ZIP10 and ZIP12 mRNAs were not detected in any tissue and ZnT3 mRNA was only identified in the kidney. All other genes were expressed in all tissues but only a few gene expression patterns allowed a significant (P<.0001) fitting of broken-line regression models, indicating homeostatic regulation under the present experimental conditions. Interestingly, these genes could be subcategorized by showing significant turnarounds in their response patterns, either at ~40 or ~60 mg Zn/kg diet (P<.0001). In conclusion, the present study showed clear differences in Zn transporter gene expression in response to SZD compared to the present literature on clinical models. We recognized that certain Zn transporter genes were regulated under the present experimental conditions by two distinct homeostatic networks. For the best of our knowledge, this represents the first comprehensive screening of Zn transporter gene expression in a highly translational model to human physiology.     

Lack of in vivo DNA binding of mercaptobenzothiazole to selected tissues of the rat

In this study, the in vivo binding of 14C-labelled 2-mercaptobenzothiazole (MBT) to DNA was investigated. Male and female Fischer 344 rats were gavaged with 375 mg MBT/kg body weight and killed 8 hours later. DNA was extracted from the liver, adrenal glands, pituitary gland, pancreas, and bone marrow and the amount of radioactivity associated with the DNA was determined. Results from this study indicate that MBT does not significantly bind to DNA from any of the tissues examined. CBI values for liver for the 3 methods of purification were -1-3 which are on the low end of the covalent binding index. The CBI values for the other tissues were always less than 1. Other chemicals with similar CBI values include estrone and diethylstilbesterol. Strong hepatocarcinogens such as dimethylnitrosamine and aflatoxin have CBI values ranging from 6000 to greater than 20000.     

The effects of severe zinc deficiency on intestinal amino acid losses in the rat

To determine whether intestinal amino acid losses might occur during zinc deficiency, labeled aminoisobutyric acid was given parenterally to zinc deficient rats and to appropriate zinc-sufficient controls. After 24 hours, the aminoisobutyric acid loss into the intestinal lumen was measured by in situ perfusion of isolated intestinal segments under conditions of either net water absorption or water secretion. Net amino acid losses were larger in the jejunum of the zinc deficient rats and losses were exacerbated during net water secretion in the jejunum and colon segments. The contribution of amino acid losses to fecal nitrogen, particularly during osmotic diarrhea, may be important in the growth retardation of zinc deficiency. Further, these alterations may indicate defective enterocyte transport functions during severe deficiency.     

Biological consequences of zinc deficiency in the pathomechanisms of selected diseases

From many points of view, zinc is one of the most important trace elements in biological systems. Many articles describe the well-known role of this metal in human physiology and pathophysiology, but in the related literature, there is a lack of current and reliable reviews of the role of zinc deficiency in many diseases. In this article, we describe the role of zinc deficiency in the oxidative stress control, immune response, proliferation, and pathogenesis and pathophysiology of selected diseases such as depression, cardiovascular diseases, diabetes mellitus, Alzheimer’s disease, and Wilson’s disease.     

Phosphorylation of rat brain calmodulin in vivo and in vitro

After injection of [32p]orthophosphate into the third ventricle of rat brain, calmodulin(CaM) was prepared from soluble(S2) and particulate(P2) fractions of the whole brain and analyzed by SDS-PAGE in the presence or absence of Ca2+ followed by autoradiography. CaM from both fractions(S2 and P2) was significantly phosphorylated by endogenous protein kinase(s) of rat brain. The incorporation of radioactive phosphate into membrane-bound CaM from the P2 fraction was much higher than that of soluble CaM from the S2 fraction. CaM was phosphorylated in vitro by casein kinase 2 but not by casein kinase 1 or by cyclic AMP-dependent protein kinase, suggesting that casein kinase 2 may be, at least in part, responsible for the phosphorylation of CaM even in vivo.     

Dietary zinc deficiency decreases plasma concentrations of vitamin E

Experiments were conducted to examine the effects of dietary zinc (Zn) upon plasma vitamin E (E) concentrations to test the hypothesis that there may be a significant dietary interaction between these two nutrients. Weanling female Sprague-Dawley rats were fed diets that were (i) Zn-deficient (less than 0.9 micrograms Zn/g diet) ad libitum; (ii) Zn-adequate (50.9 micrograms Zn/g diet), pair-fed to the Zn-deficient group; and (iii) Zn-adequate (50.9 micrograms Zn/g diet) ad libitum. Plasma E in Zn-deficient animals (4.02 +/- 1.20 micrograms/ml) was significantly reduced (P less than or equal to 0.05) compared with results in both Zn-adequate pair-fed (9.21 +/- 0.70 micrograms/ml) and Zn-adequate ad libitum-fed (9.47 +/- 0.90 micrograms/ml) animals. Zn deficiency in this model system also resulted in significant (P less than or equal to 0.05) reductions in femur and plasma Zn concentrations as well as in plasma retinol, plasma triglyceride, and plasma cholesterol concentrations. Plasma albumin and total plasma protein concentrations were normal in Zn-deficient animals. With dietary Zn deficiency, the decrease in plasma E appeared to be out of proportion to associated decreases in plasma triglyceride and plasma cholesterol concentrations. Since E is associated with plasma lipoproteins, these data suggest that lipid and/or E malabsorption may be a consequence of Zn deficiency. In response to increased dietary intake of E, increments of plasma E were lower in Zn-depleted than in Zn-adequate, pair-fed animals. These findings suggest that dietary Zn deficiency possibly may increase the nutritional requirement for E necessary to maintain adequate plasma concentrations.     

In vivo and in vitro effects of beta-endorphin on glucose metabolism in the rat

The effects of beta-endorphin (beta-Ep) on plasma glucose levels in rats and on glucose metabolism in isolated rat liver cells were examined. Intravenous injection of beta-Ep (5 micrograms/100 g BW) into ether-anaesthetized rats resulted in prompt and sustained hyperglycaemia with increases in the plasma glucagon and somatostatin levels and decrease in the plasma insulin level. When liver cells isolated from fed rats were incubated in the presence of beta-Ep at concentrations of 6 X 10(-8) M to 6 X 10(-7) M, glucose release into the medium increased within 15 min in a dose-related manner. Time course experiments showed that beta-Ep increased the level of cyclic AMP within 3 min. Significant increase in gluconeogenesis in liver cells isolated from fasted rats was also observed on addition of 10(-7) M beta-Ep in the presence of 10 mM L-lactate. These results suggest that the hyperglycaemia induced by beta-Ep may be caused, at least in part, by the effects of beta-Ep on releases of pancreatic hormones and glucose production in liver cells.     

Localization of lipoprotein lipase mRNA in selected rat tissues

Measurements of enzymatic activity have demonstrated that lipoprotein lipase (LPL), the principal enzyme responsible for hydrolysis of circulating triglyceride, is present in a number of tissues including brain, kidney, and adrenal gland. To determine the sites of synthesis of LPL in these tissues, in situ hybridization studies were performed using a non-sense 35S-labeled RNA probe produced from a 624-bp mouse LPL cDNA fragment. Control studies were performed with a sense RNA strand. Using 5-10-micron sections of 5-day-old rat brain, strong hybridization was found in pyramidal neurons of the hippocampus. Positive hybridization, indicating the presence of LPL mRNA, was also found in brain cortex and in the intermediate lobe of adult rat pituitary gland. Specific areas of adrenal and kidney medulla showed hybridization with the probe. LPL mRNA is, therefore, present in a number of specific regions of the body. LPL in these areas may not be important in regulating circulating levels of lipoproteins, but may be essential for cellular uptake, binding, and transfer of free fatty acids or other lipophilic substances.     

The effects of zinc deficiency on pancreatic carboxypeptidase activity and protein digestion and absorption in the rat

1. Proteolytic enzyme activities were examined in the pancreas of zinc-deficient and control rats. 2. No change was detected in trypsin-plus-chymotrypsin activity. 3. Carboxypeptidase activity was appreciably lowered in zinc deficiency and returned rapidly to normal on zinc therapy. 4. In experiments in which U-¹⁴C-labelled Chlorella protein was fed no evidence was obtained which suggested that the reduction in carboxypeptidase activity had limited the rate of protein digestion or absorption. 5. The specific activity of pancreatic protein synthesized during these experiments was appreciably lower in zinc-deficient than in control rats. 6. A higher proportion of the total activity present, in each organ examined, was in the non-protein fraction in zinc-deficient rats.     

In vivo and in vitro effects of aluminum treatment on rat liver mitochondrial function

This study examines the effect on mitochondrial respiration and permeability of in vivo and in vitro aluminium (Al) exposure. Rats were treated intraperitoneally with AlCl₃ to achieve serum and liver Al concentrations comparable to those seen in Al-related disorders. Mitochondria isolated from Al-treated rats had higher (p<0.01) Al concentration, lower (p<0.05) state 3 respiration, respiratory control (RCR), and ADP/O ratio (succinate substrate), and greater passive swelling in 100 mM KCl or 200 mM NH₄NO₃ than controls. The in vitro addition of Al (0–180 μM) to mitochondria from normal rats also decreased (p<0.01) state 3 respiration, RCR, and ADP/O and stimulated passive swelling in KCl and NH₄NO₃ at 42–180 μM Al. These studies show that Al depresses mitochondrial energy metabolism and increases membrane permeability. The toxicity associated with Al may be related to its effect on mitochondria.