Prolactin directly stimulated the solvent drag-induced calcium transport in the duodenum of female rats

Prolactin has been reported to stimulate the calcium absorption of the duodenum where three components of the active calcium transport, namely transcellular active, voltage-dependent and solvent drag-induced calcium transport, were identified. It was known that the transcellular active, but not the voltage-dependent, duodenal calcium transport was directly stimulated by prolactin. The present study thus aimed to evaluate the direct action of prolactin on the solvent drag-induced duodenal calcium transport by using the Ussing chamber technique. The jejunum was used as a reference for the existence of solvent drag and the widening of tight junction induced by cytochalasin E. Results showed that the solvent drag-induced calcium transport existed in both intestinal segments, but the magnitude was significantly greater in the duodenum (29.27+/-2.27 vs. 17.31+/-1.65 nmol h(-1) cm(-2), P<0.001). We further demonstrated that 200, 600 and 800, but not 1000 ng/ml, prolactin significantly promoted the solvent drag-induced duodenal calcium transport in a dose-response manner, i.e. from the control value of (nmol h(-1) cm(-2)) 24.31+/-2.36 to 45.42+/-3.47 (P<0.01), 63.82+/-5.28 (P<0.001) and 53.93+/-5.41 (P<0.01), respectively. However, prolactin did not manifest any effect on the jejunum. Because the paracellular transport was suggested to be size-selective as well as charge-selective, further experiments were designed to evaluate the mechanism by which prolactin stimulated the solvent drag-induced calcium transport. The duodenum was exposed to 20 microM cytochalasin E, 600 ng/ml prolactin or the combination of both in the presence of a paracellular marker 3H-mannitol, while the jejunum was a positive reference. The results showed that, in the jejunum, cytochalasin E alone and cytochalasin E plus prolactin significantly increased the mannitol fluxes from (micromol h(-1) cm(-2)) 0.29+/-0.04 to 0.49+/-0.03 (P<0.05) and 0.48+/-0.05 (P<0.05), respectively, while having no effect on the calcium fluxes. Prolactin alone had no effect on the jejunal calcium flux. In the duodenum, neither mannitol nor calcium fluxes were enhanced by cytochalasin E, however, prolactin still increased the solvent drag-induced calcium flux from 27.74+/-2.41 to 51.03+/-4.35 nmol h(-1) cm(-2) (P<0.001). It was concluded that prolactin directly stimulated the solvent drag-induced duodenal calcium transport in a dose-response and biphasic manner without the widening of tight junction.     

Prolactin stimulates the L-type calcium channel-mediated transepithelial calcium transport in the duodenum of male rats

Elevated plasma levels of prolactin (PRL) have been reported in several physiological and pathological conditions, such as lactation, prolactinoma, and dopaminergic antipsychotic drug uses. Although PRL is a calcium-regulating hormone that stimulates intestinal calcium absorption in lactating rats, whether PRL is capable of stimulating calcium absorption in male rats has been elusive. Herein, the transepithelial calcium transport and electrical characteristics were determined in ex vivo duodenal tissues of male rats by Ussing chamber technique. We found that PRL receptors were abundantly present in the basolateral membrane of the duodenal epithelial cells. PRL (200–800 ng/mL) markedly increased the active duodenal calcium transport in a dose-dependent fashion without effect on the transepithelial resistance. The PRL-enhanced active duodenal calcium transport was completely abolished by L-type calcium channel blocker (nifedipine) as well as inhibitors of the major basolateral calcium transporters, namely plasma membrane Ca²⁺-ATPase and Na⁺/Ca²⁺ exchanger. Several intracellular mediators, such as JAK2, MEK, PI3K and Src kinase, were involved in the PRL-enhanced transcellular calcium transport. Moreover, PRL also stimulated the paracellular calcium transport in the duodenum of male rats in a PI3K-dependent manner. In conclusion, PRL appeared to be a calcium-regulating hormone in male rats by enhancing the L-type calcium channel-mediated transcellular and the paracellular passive duodenal calcium transport. This phenomenon could help restrict or alleviate negative calcium balance and osteoporosis that often accompany hyperprolactinemia in male patients.     

Chronic metabolic acidosis stimulated transcellular and solvent drag-induced calcium transport in the duodenum of female rats

Chronic metabolic acidosis results in a negative calcium balance as a result of bone resorption and renal calcium loss. However, reports on the changes in intestinal calcium transport have been controversial. The present investigation therefore aimed to study the effects of chronic metabolic acidosis induced by 1.5% NH(4)Cl administration on the three components of duodenal calcium transport, namely, solvent drag-induced, transcellular active, and passive paracellular components, in rats using an in vitro Ussing chamber technique. The relative mRNA expression of genes related to duodenal calcium transport was also determined. We found that 21-day chronic metabolic acidosis stimulated solvent drag-induced and transcellular active duodenal calcium transport but not passive paracellular calcium transport. Our results further demonstrated that an acute direct exposure to serosal acidic pH, in contrast, decreased solvent drag-induced calcium transport in a pH-dependent fashion but had no effect on transcellular active calcium transport. Neither the transepithelial resistance nor duodenal permeability to Na(+), Cl(-), and Ca(2+) via the passive paracellular pathway were altered by chronic metabolic acidosis, suggesting that widening of the tight junction and changes in the charge-selective property of the tight junction did not occur. Thus the enhanced duodenal calcium transport observed in chronic metabolic acidosis could have resulted from a long-term adaptation, possibly at the molecular level. RT-PCR study revealed that chronic metabolic acidosis significantly increased the relative mRNA expression of duodenal genes associated with solvent drag-induced transport, i.e., the beta(1)-subunit of Na(+)-K(+)-ATPase, zonula occludens-1, occludin, and claudin-3, and with transcellular active transport, i.e., transient receptor potential vanilloid family Ca(2+) channels 5 and 6 and plasma membrane Ca(2+)-ATPase isoform 1b. Total plasma calcium and free ionized calcium and magnesium concentrations were also increased, whereas serum parathyroid hormone and 1alpha,25-dihydroxyvitamin D(3) levels were not changed. The results indicated that 21-day chronic metabolic acidosis affected the calcium metabolism in rats partly through enhancing the mRNA expression of crucial duodenal genes involved in calcium absorption, thereby stimulating solvent drag-induced and transcellular active calcium transport in the duodenum.     

Long-term prolactin exposure differentially stimulated the transcellular and solvent drag-induced calcium transport in the duodenum of ovariectomized rats

Prolactin, having been shown to stimulate transcellular active and solvent drag-induced calcium transport in the duodenum of female rats, was postulated to improve duodenal calcium transport in estrogen-deficient rats. The aim of the present study was, therefore, to demonstrate the effects of long-term prolactin exposure produced by anterior pituitary (AP) transplantation on the duodenal calcium transport in young (9-week-old) and adult (22-week-old) ovariectomized rats. We found that ovariectomy did not alter the transcellular active duodenal calcium transport in young and adult rats fed normal calcium diet (1.0% w/w Ca) but decreased the solvent drag-induced duodenal calcium transport from 75.50 +/- 10.12 to 55.75 +/- 4.77 nmol.hr(-1).cm(-2) (P < 0.05) only in adult rats. Long-term prolactin exposure stimulated the transcellular active calcium transport in young and adult AP-grafted ovariectomized rats fed with normal calcium diet by more than 2-fold from 7.56 +/- 0.79 to 16.54 +/- 2.05 (P < 0.001) and 9.78 +/- 0.72 to 15.99 +/- 1.75 (P < 0.001) nmol.hr(-1).cm(-2), respectively. However, only the solvent drag-induced duodenal calcium transport in young rats was enhanced by prolactin from 95.51 +/- 10.64 to 163.20 +/- 18.03 nmol.hr(-1).cm(-2) (P < 0.001) whereas that in adult rats still showed a decreased flux from 75.50 +/- 10.12 to 47.77 +/- 5.42 nmol.hr(-1).cm(-2) (P < 0.05). Because oral calcium supplement has been widely used to improve calcium balance in estrogen-deficient animals, the effect of a high-calcium diet (2.0% w/w Ca) was also investigated. The results showed that stimulatory action of long-term prolactin on the transcellular active duodenal calcium transport in both young and adult rats was diminished after being fed a high-calcium diet. The same diet also abolished prolactin-enhanced solvent drag-induced duodenal calcium transport in young and further decreased that in adult AP-grafted ovariectomized rats. We concluded that the solvent drag-induced duodenal calcium transport in adult rats was decreased after ovariectomy. Long-term prolactin exposure stimulated the transcellular active duodenal calcium transport in both young and adult rats whereas enhancing the solvent drag-induced duodenal calcium transport only in young rats. Effects of prolactin were abolished by a high-calcium diet.     

Regulation of duodenal insulin-like growth factor I and active calcium transport by ovariectomy in female rats.

There is a significant body of data that supports the concept that reproductive hormones in females have effects on duodenal calcium transport that are not mediated via altered circulating concentrations of 1,25-dihydroxyvitamin D (1,25(OH)2D). Previously, we have shown parallel alterations in duodenal Ca transport and longitudinal bone growth rate in sexually maturing female rats in response to ovariectomy and estradiol (E) treatment of ovariectomized (OVX) rats (OVX+E) without any change in circulating levels of 1,25(OH)2D or parathyroid hormone. Results are presented here from experiments designed to: (i) further explore the relationship between 1,25(OH)2D and ovarian status in the regulation of duodenal calcium transport, and (ii) determine whether OVX and E replacement alter circulating and duodenal levels of insulin-like growth factor I (IGF-I) that might be related to effects on Ca transport. Growth hormone, which has been shown to affect intestinal Ca absorption and vitamin D metabolism, is thought to act indirectly by stimulating IGF-I. Six-week-old female rats were OVX, given estradiol implants (OVX+E), and fed a diet containing either 0.5% or 0.1% Ca for 3 weeks. In both diet groups, the OVX animals exhibited a higher level of Ca transport, as measured by the everted gut sac method, than either the intact controls or the OVX+E group; there was no difference in calcium transport between the different diet groups. Although there was no difference in circulating levels of 1,25(OH)2D among the intact, OVX, and OVX+E groups fed either diet, animals fed the 0.1% Ca diet had higher circulating levels of 1,25(OH)2D than those fed the 0.5% Ca diet. There was no difference in duodenal levels of calbindin9K among intact, OVX, and OVX+E animals in either diet group, although the animals fed the 0.1% Ca diet had higher levels of calbindin9K than the animals fed the 0.5% Ca diet. In animals fed the 0.5% Ca diet, OVX resulted in elevated serum and duodenal levels of IGF-1, as compared with intact and OVX+E animals on the same diet. In animals fed the 0.1% Ca diet, there was no elevation of IGF-I in the OVX group relative to intact and OVX+E animals. These results lend additional support to the concept that alterations in duodenal active calcium transport that occur with alterations in ovarian hormones are not mediated by changes in serum levels of 1,25(OH)2D, but may be related to some factor related to growth, possibly IGF-I.     

Functional organization of calcium stores in polarized secretory cells and transcellular calcium transport

In this short review, we will first discuss localized cytoplasmic calcium signals in pancreatic acinar cells. In the second part of the review, we will describe recently discovered polarized calcium efflux and calcium propagation through the lumen of the endoplasmic reticulum — ER (a phenomenon we have termed “calcium tunnelling”). Finally, we will present a hypothesis concerning the roles that these mechanisms could play in transcellular calcium flux.     

Prolactin is an important regulator of intestinal calcium transport

Prolactin has been shown to stimulate intestinal calcium absorption, increase bone turnover, and reduce renal calcium excretion. The small intestine, which is the sole organ supplying new calcium to the body, intensely expresses mRNAs and proteins of prolactin receptors, especially in the duodenum and jejunum, indicating the intestine as a target tissue of prolactin. A number of investigations show that prolactin is able to stimulate the intestinal calcium transport both in vitro and in vivo, whereas bromocriptine, which inhibits pituitary prolactin secretion, antagonizes its actions. In female rats, acute and long-term exposure to high prolactin levels significantly enhances the (i) transcellular active, (ii) solvent drag-induced, and (iii) passive calcium transport occurring in the small intestine. These effects are seen not only in pregnant and lactating animals, but are also observed in non-pregnant and non-lactating animals. Interestingly, young animals are more responsive to prolactin than adults. Prolactin-enhanced calcium absorption gradually diminishes with age, thus suggesting it has an age-dependent mode of action. Although prolactin's effects on calcium absorption are not directly vitamin D-dependent; a certain level of circulating vitamin D may be required for the basal expression of genes related to calcium transport. The aforementioned body of evidence supports the hypothesis that prolactin acts as a regulator of calcium homeostasis by controlling the intestinal calcium absorption. Cellular and molecular signal transductions of prolactin in the enterocytes are largely unknown, however, and still require investigation.     

The pyroantimonate reaction and transcellular transport of calcium in rat molar enamel organs

Summary The distribution of calcium in the cells of the enamel organ of developing rat molar tooth germs was studied by the pyroantimonate method. It was found that there was a specific localization to the inner leaflet of the plasma membrane of both secretory and maturation phase ameloblasts. This information can be used to support the model for transcellular transport of calcium involving membrane fluidity, with phosphatidylserine as a carrier (Reith 1983). It can also support an alternative model involving movement of calcium ions over a surface of acidic phospholipids on the inner leaflet of the plasma membrane, without involving the necessity for membrane fluidity.Deceased 9 May 1985     

The pyroantimonate reaction and transcellular transport of calcium in rat molar enamel organs

The distribution of calcium in the cells of the enamel organ of developing rat molar tooth germs was studied by the pyroantimonate method. It was found that there was a specific localization to the inner leaflet of the plasma membrane of both secretory and maturation phase ameloblasts. This information can be used to support the model for transcellular transport of calcium involving membrane fluidity, with phosphatidylserine as a carrier (Reith 1983). It can also support an alternative model involving movement of calcium ions over a surface of acidic phospholipids on the inner leaflet of the plasma membrane, without involving the necessity for membrane fluidity.     

Modeling of transcellular Ca transport in rat duodenum points to coexistence of two mechanisms of apical entry

Employing realistic parameters, we have demonstrated that a relatively simple mathematical model can reproduce key features of steady-state Ca2+ transport with the assumption of two mechanisms of Ca2+ entry: a channel-like flux and a carrier-mediated transport. At low luminal [Ca2+] (1-5 mM), facilitated entry dominates and saturates with Km = 0.4 mM. At luminal [Ca2+] of tens of millimolar, apical permeability is dominated by the channel flux that in turn is regulated by cytosolic Ca2+. The model reproduces the linear relationship between maximum Ca2+ transport rate and intestinal calbindin D9K (CaBP) content. At luminal [Ca2+] > 50 mM, local sensitivity analysis shows transcellular transport to be most sensitive to variations in CaBP. At low luminal [Ca2+], transport becomes sensitive to apical entry regulation. The simulations have been run within the Virtual Cell modeling environment, yielding the time course of external Ca2+ and spatiotemporal distributions of both intracellular Ca2+ and CaBP. Coexistence of two apical entry mechanisms accords with the properties of the duodenal Ca2+ transport protein CaT1 and the epithelial Ca2+ channel ECaC.     

Prolactin stimulates precursor cells in the adult mouse hippocampus

In the search for ways to combat degenerative neurological disorders, neurogenesis-stimulating factors are proving to be a promising area of research. In this study, we show that the hormonal factor prolactin (PRL) can activate a pool of latent precursor cells in the adult mouse hippocampus. Using an in vitro neurosphere assay, we found that the addition of exogenous PRL to primary adult hippocampal cells resulted in an approximate 50% increase in neurosphere number. In addition, direct infusion of PRL into the adult dentate gyrus also resulted in a significant increase in neurosphere number. Together these data indicate that exogenous PRL can increase hippocampal precursor numbers both in vitro and in vivo. Conversely, PRL null mice showed a significant reduction (approximately 80%) in the number of hippocampal-derived neurospheres. Interestingly, no deficit in precursor proliferation was observed in vivo, indicating that in this situation other niche factors can compensate for a loss in PRL. The PRL loss resulted in learning and memory deficits in the PRL null mice, as indicated by significant deficits in the standard behavioral tests requiring input from the hippocampus. This behavioral deficit was rescued by direct infusion of recombinant PRL into the hippocampus, indicating that a lack of PRL in the adult mouse hippocampus can be correlated with impaired learning and memory.     

Glucose 1-phosphate increases active transport of calcium in intestine

Active calcium transport in intestine is essential for serum calcium homeostasis as well as for bone formation. It is well recognized that vitamin D is a major, if not sole, stimulator of intestinal calcium transport activity in mammals. Besides vitamin D, endogenous glucose 1-phosphate (G1P) affects calcium transport activity in some microorganisms. In this study, we investigated whether G1P affects intestinal calcium transport activity in mammals as well. Of several glycolytic intermediates, G1P was the sole sugar compound in stimulating intestinal calcium uptake in Caco-2 cells. G1P stimulated net calcium influx and expression of calbindin D9K protein in rat intestine, through an active transport mechanism. Calcium uptake in G1P-supplemented rats was greater than that in the control rats fed a diet containing adequate vitamin D3. Bone mineral density (BMD) of aged rat femoral metaphysis and diaphysis was also increased by feeding the G1P diet. G1P did not affect serum levels of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] at all. These results suggest that exogenously applied G1P stimulates active transport of calcium in intestine, independent of vitamin D, leading to an increase of BMD.     

Marker enzymes in rat liver vesicles involved in transcellular transport

In order to label the vesicles involved in transcellular transfer (transcytosis) through hepatocytes, polymeric IgA (pIgA) was conjugated to horseradish peroxidase (HRP) and injected into rats. The endosomes containing this ligand at 10 or 20 min after injection were isolated by the diaminobenzidine-induced density-shift procedure and their content in various marker enzymes was measured. The endosomes carrying pIgA-HRP 10 min after injection contained only traces of 5'-nucleotidase and low amounts of alkaline phosphodiesterase I. The estimated marker enzyme content is similar to that observed for the particles containing galactosylated bovine serum albumin conjugated to HRP, a ligand degraded in lysosomes. However, 20 min after injection, the transcytotic endosomes showed a marked enrichment in 5'-nucleotidase and especially in alkaline phosphodiesterase I. The results confirm the heterogeneity of rat liver endosomes and substantiate the concept of distinct endosomal compartments.