Luminal Ca Concentration Research Articles

Calcium transport in bovine rumen epithelium as affected by luminal Ca concentrations and Ca sources.

The quantitative role of different segments of the gastrointestinal tract for Ca absorption, the respective mechanisms, and their regulation are not fully identified for ruminants, that is, cattle. In different in vitro experiments the forestomach wall has been demonstrated to be a major site for active Ca absorption in sheep and goats. In order to further clarify the role of the bovine rumen for Ca transport with special attention to luminal Ca concentrations, its ionic form, and pH, electrophysiological and unidirectional flux rate measurements were performed with isolated bovine rumen epithelial tissues. For Ca flux studies (Jms, Jsm) in vitro Ussing chamber technique was applied. Standard RT-PCR method was used to characterize TRPV6 and PMCA1 as potential contributors to transepithelial active Ca transport. At Ca concentrations of 1.2 mmol L−1 on both sides of the tissues, Jms were higher than Jsm resulting under some conditions in significant Ca net flux rates (Jnet), indicating the presence of active Ca transport. In the absence of an electrical gradient, Jnet could significantly be stimulated in the presence of luminal short-chain fatty acids (SCFAs). Increasing the luminal Ca concentrations up to 11.2 mmol L−1 resulted in significant increases in Jms without influencing Jsm. Providing Ca in its form as respective chloride, formate, or propionate salts there was no significant effect on Jms. No transcripts specific for Ca channel TRPV6 could be demonstrated. Our results indicate different mechanisms for Ca absorption in bovine rumen as compared with those usually described for the small intestines.

Divalent cation‐dependent 3H‐L‐leucine transport in lobster intestine

The crustacean intestine displays a diversity of transport mechanisms for the transport of dietary substances, including metal ions and amino acids. The intestine of the American lobster Homarus americanus was isolated and perfused in vitro with a physiological saline. Transmural 20 μM mucosa to serosa (MS) 3H‐L‐leucine fluxes across the intestine were a hyperbolic function of luminal Ca concentration. Ca‐dependent 3H‐L‐leucine transport exhibited low Km (high affinity) and low Jmax compared to kinetic constants when both Ca and Zn were simultaneously present, and under these conditions amino acid transport Jmax was doubled. MS flux of 3H‐L‐leucine was a hyperbolic function of L‐leucine and was stimulated by the addition of 20 μM luminal Mn. In the presence of Mn alone, 3H‐L‐leucine transport showed an increase in Jmax, but no effect on L‐leucine influx Km. With the addition of Ca, the Jmax decreased, but Ca had no effect on L‐leucine influx Km, suggesting that Ca may be acting as an allosteric inhibitor of metal‐stimulated amino acid transport. In other experiments calcium was a non‐competitive inhibitor of manganese activation of 3H‐L‐leucine transport, apparently binding to a site other than the metal binding site. Ongoing experiments are researching the nature of Ca inhibition of metal stimulated amino acid transport.Agriculture and Food Research Initiative Competitive USDA Grant no. 2010–65206‐20617.

Luminal calcium concentration controls intestinal calcium absorption by modification of intestinal alkaline phosphatase activity

Intestinal alkaline phosphatase (IAP) is a brush-border phosphomonoesterase. Its location suggests an involvement in the uptake of nutrients, but its role has not yet been defined. IAP expression parallels that of other proteins involved in Ca absorption under vitamin D stimulation. Experiments carried out in vitro with purified IAP have demonstrated an interaction between Ca and IAP. The gut is prepared to face different levels of Ca intake over time, but high Ca intake in a situation of a low-Ca diet over time would cause excessive entry of Ca into the enterocytes. The presence of a mechanism to block Ca entry and to avoid possible adverse effects is thus predictable. Thus, in the present study, Sprague-Dawley rats were fed with different amounts of Ca in the diet (0.2, 1 and 2 g%), and the percentage of Ca absorption (%Ca) in the presence and absence of L-phenylalanine (Phe) was calculated. The presence of Phe caused a significant increase in %Ca (52.3 (SEM 6.5) % in the presence of Phe v. 31.1 (sem 8.9) % in the absence of Phe, regardless of the amount of Ca intake; paired t test, P = 0.02). When data were analysed with respect to Ca intake, a significant difference was found only in the group with low Ca intake (paired t test, P = 0.03). Additionally, IAP activity increased significantly (ANOVA, P < 0.05) as Ca concentrations increased in the duodenal lumen. The present study provides in vivo evidence that luminal Ca concentration increases the activity of IAP and simultaneously decreases %Ca, acting as a minute-to-minute regulatory mechanism of Ca entry.

Axotomy depletes intracellular calcium stores in primary sensory neurons.

The cellular mechanisms of neuropathic pain are inadequately understood. Previous investigations have revealed disrupted Ca signaling in primary sensory neurons after injury. The authors examined the effect of injury on intracellular Ca stores of the endoplasmic reticulum, which critically regulate the Ca signal and neuronal function. Intracellular Ca levels were measured with Fura-2 or mag-Fura-2 microfluorometry in axotomized fifth lumbar (L5) dorsal root ganglion neurons and adjacent L4 neurons isolated from hyperalgesic rats after L5 spinal nerve ligation, compared to neurons from control animals. Endoplasmic reticulum Ca stores released by the ryanodine-receptor agonist caffeine decreased by 46% in axotomized small neurons. This effect persisted in Ca-free bath solution, which removes the contribution of store-operated membrane Ca channels, and after blockade of the mitochondrial, sarco-endoplasmic Ca-ATPase and the plasma membrane Ca ATPase pathways. Ca released by the sarco-endoplasmic Ca-ATPase blocker thapsigargin and by the Ca-ionophore ionomycin was also diminished by 25% and 41%, respectively. In contrast to control neurons, Ca stores in axotomized neurons were not expanded by neuronal activation by K depolarization, and the proportionate rate of refilling by sarco-endoplasmic Ca-ATPase was normal. Luminal Ca concentration was also reduced by 38% in axotomized neurons in permeabilized neurons. The adjacent neurons of the L4 dorsal root ganglia showed modest and inconsistent changes after L5 spinal nerve ligation. Painful nerve injury leads to diminished releasable endoplasmic reticulum Ca stores and a reduced luminal Ca concentration. Depletion of Ca stores may contribute to the pathogenesis of neuropathic pain.

Tricyclic antidepressant amitriptyline alters sarcoplasmic reticulum calcium handling in ventricular myocytes

Tricyclic antidepressants such as amitriptyline (AMT) have been reported to have adverse side effects on cardiac performance. AMT effects on Ca handling in ventricular myocytes, however, are not well understood. Therefore, we investigated AMT action on sarcoplasmic reticulum (SR) Ca release in ventricular myocytes, ryanodine receptor (RyR) activity, and Ca uptake by SR microsomes. In permeabilized myocytes, AMT transiently increased free luminal Ca concentration ([Ca]) followed by marked depletion. AMT (10 microM) caused a rapid and a transient increase of Ca spark frequency, followed by a significant suppression of spark activity. The latter was associated with a decrease of Ca spark amplitude and SR Ca load to 87 and 60%, respectively. AMT (10 microM) completely abolished propagation of spontaneous Ca waves. Higher concentrations of AMT (0.1-1 mM) evoked SR Ca release reminiscent of the effect of caffeine (20 mM) and caused almost complete depletion of SR Ca content. Studies on single calsequestrin-free RyR channels revealed that AMT increased the mean open time and open probability (Po) in a dose-dependent fashion (dissociation constant = 4.2 microM). High concentrations of AMT (> 25 microM) evoked frequent long openings with Po reaching very high levels (> 0.70). In studies with cardiac SR microsomes, AMT slowed the rate of ATP-dependent Ca uptake. We conclude that AMT affects SR Ca handling in ventricular myocytes by multiple mechanisms, including direct stimulation of RyRs and inhibition of SR Ca uptake. These effects could contribute to AMT cardiotoxicity.

Maturation of Calcium Transport in the Rat Small and Large Intestine

Using an in vivo perfusion technique we compared transport of calcium in segments of the proximal (duodenum + jejunum) and distal (ileum) small intestine and in the cecum + colon of suckling (2-week old), weanling (3-week-old) and adolescent (6-week-old) rats. The concentration of calcium (Ca) in the isotonic sodium chloride solution perfused through the intestinal segments was either below (0.4 and 1.4 mmoles/1) or above (3.2 mmoles/1) serum ionized calcium. In the suckling rat, in all three intestinal segments, the relationship between transport rate (mumoles/hr per g wet weight) and luminal Ca concentration was linear. Net secretion occurred when luminal Ca concentration was below serum ionized Ca, and net absorption at 3.2 mmoles/l luminal Ca. In the weanling rats the relationship between luminal Ca concentration and transport rate was curvilinear for all segments. In the adolescent rats the relationship was curvilinear in the proximal segment and linear in the distal segment and in the cecum + colon. Rates of transport in segments of the suckling rats were several fold greater than corresponding rates in the adolescent rats. The change in the relationship between transport rate and luminal Ca suggested that the mode of transport of Ca changes at the time of weaning from mainly a passive mechanism in the suckling period to mainly a carrier mediated mechanism in the adolescent rats.

425 INTESTINAL MATURATION: CALCIUM (Ca) TRANSPORT

Our previous study suggested that the permeability of the intestinal mucosa (PIM) to Ca was greater in suckling (S) than in adolescent (A) rats (Pediat. Res. 11:414, 1977). Definitive studies comparing PIM to Ca during development are not available. We compared transport of Ca in the proximal intestine of S (2 week old), W (3 week old) and A (6 week old) rats, using an in vivo recirculation perfusion technique. The perfusates were isotonic NaCl solution containing either 0.4, 1.6 or 3.4 mmoles of CaCl2/1 and Phenol Red 20 mg/1. Ca transport rates (mean±S.E.; μmoles/hr per g. wet wt.) from the different solutions were: 0.4 mmole/1 (S, -32.3±6.5; A, -0.1±0.2; p<.001); 1.6 mmole/l (S, -14.8±2.3; A, 1.8±0.4; p<.005); 3.4 mmole/1 (S, 27.6±6.0; A, 3.3±0.9; p<.005). Transport (T) rates in W rats were intermediate between those of S and A rats (-4.5±1.6, 5.1±1.3, 19.0±1.5 respectively, from solutions containing 0.4, 1.6 or 3.4 mmole Ca/1). The 8 fold greater net absorption (+T) rate at the high, and much greater net secretion (-T) rate at the low luminal Ca concentrations in S than A rats indicated that PIM to Ca was greater in S than A rats. The reason for the age difference is not known. However, the process of maturation appears to decrease the PIM to Ca, which continues into the post-weaning age. If the age differences in Ca transport is also present in humans, young infants, especially those with low birth weight, could be at risk of losing relatively large amounts of Ca into the intestinal lumen when dietary Ca intake stops abruptly.