Search

The lactogenic hormone prolactin (PRL) directly regulates osteoblast functions in vitro and modulates bone remodeling in nulliparous rats, but its osteoregulatory roles in pregnant and lactating rats with physiological hyperprolactinemia remained unclear. Herein, bone changes were investigated in rats treated with bromocriptine (Bromo), an inhibitor of pituitary PRL release, or Bromo+PRL at different reproductive phases, from mid-pregnancy to late lactation. PRL receptors were strongly expressed in osteoblasts lining bone trabeculae, indicating bone as a target of PRL actions. By using dual energy X-ray absorptiometry, we found a significant increase in bone mineral density in the femora and vertebrae of pregnant rats. Such pregnancy-induced bone gain was, however, PRL independent and may have resulted from the increased cortical thickness. Bone trabeculae were modestly changed during pregnancy as evaluated by bone histomorphometry. On the other hand, lactating rats, especially in late lactation, showed massive bone loss in bone trabeculae but not in cortical shells. Further study in Bromo- and Bromo+PRL-treated rats suggested that PRL contributed to decreases in trabecular bone volume and number and increases in trabecular separation and eroded surface, as well as a paradoxical increase in bone formation rate in late lactation. Uncoupling of trabecular bone formation and resorption was evident in lactating rats, with the latter being predominant. In conclusion, pregnancy mainly induced cortical bone gain, whereas lactation led to trabecular bone loss in both long bones and vertebrae. Although PRL was not responsible for the pregnancy-induced bone gain, it was an important regulator of bone modeling during lactation. bone histomorphometry; hyperprolactinemia; ion chromatography; osteopenia; uncoupling doi: 10.1152/ajpendo.00134.2010.

During pregnancy and lactation, the enhanced intestinal [Ca.sup.2+] absorption serves to provide [Ca.sup.2+] for fetal development and lactogenesis; however, the responsible hormone and its mechanisms remain elusive. We elucidated herein that prolactin (PRL) markedly stimulated the transcellular and paracellular [Ca.sup.2+] transport in the duodenum of pregnant and lactating rats as well as in Caco-2 monolayer in a two-step manner. Specifically, a long-term exposure to PRL in pregnancy and lactation induced an adaptation in duodenal cells at genomic levels by upregulating the expression of genes related to transcellular transport, e.g., TRPV5/6 and calbindin-[D.sub.9k], and the paracellular transport, e.g., claudin-3, thereby raising [Ca.sup.2+] absorption rate to a new 'baseline' (Step 1). During suckling, PRL surge further increased [Ca.sup.2+] absorption to a higher level (Step 2) in a nongenomic manner to match [Ca.sup.2+] loss in milk. PRL-enhanced apical [Ca.sup.2+] uptake was responsible for the increased transcellular transport, whereas PRL-enhanced paracellular transport required claudin-15, which regulated epithelial cation selectivity and paracellular [Ca.sup.2+] movement. Such nongenomic PRL actions were mediated by phosphoinositide 3-kinase, protein kinase C, and RhoA-associated coiled-coil-forming kinase pathways. In conclusion, two-step stimulation of intestinal [Ca.sup.2+] absorption resulted from long-term PRL exposure, which upregulated [Ca.sup.2+] transporter genes to elevate the transport baseline, and the suckling-induced transient PRL surge, which further increased [Ca.sup.2+] transport to the maximal capacity. The present findings also suggested that [Ca.sup.2+] supplementation at 15-30 min prior to breastfeeding may best benefit the lactating mother, since more [Ca.sup.2+] could be absorbed as a result of the suckling-induced PRL surge. calcium transport; lactating rats; pituitary transplantation; pregnancy; small interfering RNA

Previous investigations suggested that prolactin (PRL) stimulated the intestinal calcium absorption through phosphoinositide 3-kinase (PI3K), protein kinase C (PKC), and RhoA-associated coiled-coil forming kinase (ROCK) signaling pathways. However, little was known regarding its detailed mechanisms for the stimulation of transcellular and voltage-dependent paracellular calcium transport. By using Ussing chamber technique, we found that the PRL-induced increase in the transcellular calcium flux and decrease in transepithelial resistance of intestinal-like Caco-2 monolayer were not abolished by inhibitors of gene transcription and protein biosynthesis. The PRL-stimulated transcellular calcium transport was completely inhibited by the L-type calcium channel blockers (nifedipine and verapamil) and plasma membrane [Ca.sup.2+]-ATPase (PMCA) inhibitor (trifluoperazine) as well as small interfering RNA targeting voltage-dependent L-type calcium channel [Ca.sub.v]1.3, but not TRPV6 or calbindin-[D.sub.9k]. As demonstrated by [sup.45]Ca uptake study, PI3K and PKC, but not ROCK, were essential for the PRL-enhanced apical calcium entry. In addition, PRL was unable to enhance the transcellular calcium transport after [PKC.sub.[zeta]] knockdown or exposure to inhibitors of [PKC.sub.[zeta]], but not of [PKC.sub.[alpha]], [PKC.sub.[beta]], [PKC.sub.[epsilon]], [PKC.sub.[mu]], or protein kinase A. Voltage-clamping experiments further showed that PRL markedly stimulated the voltage-dependent calcium transport and removed the paracellular rectification. Such PRL effects on paracellular transport were completely abolished by inhibitors of PI3K (LY-294002) and ROCK (Y-27632). It could be concluded that the PRL-stimulated transcellular calcium transport in Caco-2 monolayer was mediated by [Ca.sub.v]1.3 and PMCA, presumably through PI3K and [PKC.sub.[zeta]] pathways, while the enhanced voltage-dependent calcium transport occurred through PI3K and ROCK pathways. calcium uptake; protein kinase C; rectification; small interfering RNA; voltage-dependent calcium transport

Endurance impact exercise, e.g., running, is known to enhance the intestinal calcium absorption. However, nonimpact exercise, e.g., swimming, is more appropriate for osteoporotic patients with cardiovascular diseases or disorders of bone and joint, but the effect of swimming on the intestinal calcium transport was unknown. This study, therefore, aimed to investigate the transepithelial calcium transport and the expression of related genes in the intestine of rats trained to swim nonstop 1 h/day, 5 days/wk for 2 wk. We found that endurance swimming stimulated calcium transport in the duodenum, proximal jejunum, and cecum, while decreasing that in the proximal colon. Swimming affected neither the transepithelial potential difference nor resistance. As demonstrated by real-time PCR, the small intestine, especially the duodenum, responded to swimming by upregulating a number of genes related to the transcellular calcium transport, i.e., TRPV5, TRPV6, calbindin-[D.sub.9k], PMC[A.sub.1b], and NCX1, and the paracellular calcium transport, i.e., ZO-1, ZO-2, ZO-3, cingulin, occludin, and claudins, as well as nuclear receptor of 1,25[(OH).sub.2][D.sub.3]. In contrast, swimming downregulated those genes in the colon. Microarray analysis showed that swimming also altered the expression of duodenal genes related to the transport of several ions and nutrients, e.g., [Na.sup.+], [K.sup.+], [C1.sup.-], glucose, and amino acids. In conclusion, endurance swimming enhanced intestinal calcium absorption, in part, by upregulating the calcium transporter genes. The present microarray study also provided relevant information for further investigations into the intestinal nutrient and electrolyte transport during nonimpact exercise. gene ontology; microarray; nonimpact exercise; real-time polymerase chain reaction; Ussing chamber

Prolactin (PRL) was previously demonstrated to rapidly enhance calcium absorption in rat duodenum and the intestine-like Caco-2 monolayer. However, its mechanism was not completely understood. Here, we investigated nongenomic effects of PRL on the transepithelial calcium transport and paracellular permselectivity in the Caco-2 monolayer by Ussing chamber technique. PRL increased the transcellular and paracellular calcium luxes and paracellular calcium permeability within 60 min after exposure but decreased the transepithelial resistance of the monolayer. The effects of PRL could not be inhibited by RNA polymerase II inhibitor (5,6-dichloro-1-[beta]-D-ribobenzimidazole), confirming that PRL actions were nongenomic. Exposure to protein kinase C (PKC) or RhoA-associated coiled-coil forming kinase (ROCK) inhibitors (GF-109203X and Y-27632, respectively) abolished the stimulatory effect of PRL on transcellular calcium transport, whereas ROCK inhibitor, but not PKC inhibitor, diminished the PRL effect on paracellular calcium transport. Knockdown of the long isoform of PRL receptor (PRLR-L) also prevented the enhancement of calcium transport by PRL. In addition, PRL markedly increased paracellular sodium permeability and the permeability ratio of sodium to chloride, which are indicators of the paracellular charge-selective property and are known to be associated with the enhanced paracellular calcium transport. The permeability of other cations in the alkali metal series was also increased by PRL, and such increases were abolished by ROCK inhibitor. It could be concluded that PRL stimulated transepithelial calcium transport through PRLR-L and increased paracellular permeability to cations in the Caco-2 monolayer. These nongenomic actions of PRL were mediated by the PKC and ROCK signaling pathways. charge selectivity; dilution potential; paracellular transport; phosphoinositide 3-kinase; protein kinase C; long form of prolactin receptor; RhoA; short interference RNA; transcellular transport

Abstract: Hyperprolactinemia caused by physiological or pathological conditions, such as those occurring during lactation and prolactinoma, respectively, results in progressive osteopenia. The underlying mechanisms, however, are controversial. Prolactin (PRL) may [...]

In type 2 diabetes mellitus (T2DM), the decreased bone strength is often associated with hyperglycemia and bone cell insulin resistance. Since T2DM is increasingly reported in young adults, it is not known whether the effect of T2DM on bone would be different in young adolescents and aging adults. Here, we found shorter femoral and tibial lengths in 7-month, but not 13-month, Goto-Kakizaki (GK) T2DM rats as compared to wild-type rats. Bone [micro]CT analysis showed long-lasting impairment of both cortical and trabecular bones in GK rats. Although insulin treatment effectively improved hyperglycemia, it was not able to rescue trabecular BMD and cortical thickness in young adult GK rats. In conclusion, insulin treatment and alleviation of hyperglycemia did not increase BMD of osteopenic GK rats. It is likely that early prevention of insulin resistance should prevail over treatment of full-blown T2DM-related osteopathy. Keywords: Blood glucose, Bone mineral density, Diabetes mellitus, Diabetic osteopathy, Goto-Kakizaki rats, Author(s): Ratchaneevan Aeimlapa[sup.1,2], Narattaphol Charoenphandhu[sup.1,2,3], Panan Suntornsaratoon[sup.1,2], Kannikar Wongdee[sup.2,4], Wacharaporn Tiyasatkulkovit[sup.2,5], Kanchana Kengkoom[sup.6] and Nateetip Krishnamra[sup.1,2] Introduction Diabetes mellitus (DM) is a globally common non-communicable disease which has deleterious effects [...]

Prolactin (PRL) has been shown to stimulate intestinal calcium absorption but the mechanism was still unknown. This study aimed to investigate the mechanism and signaling pathway by which PRL enhanced calcium transport in the rat duodenum and Caco-2 monolayer. Both epithelia strongly expressed mRNAs and proteins of PRL receptors. Ussing chamber technique showed that the duodenal active calcium fluxes were increased by PRL in a dose-response manner with the maximal effective dose of 800 ng/ml. This response diminished after exposure to LY-294002, a phosphoinositide 3-kinase (PI3K) inhibitor. Caco-2 monolayer gave similar response to PRL with the maximal effective dose of 600 ng/ml. By nullifying the transepithelial potential difference, we showed that the voltage-dependent paracellular calcium transport did not contribute to the PRL-enhanced flux in Caco-2 monolayer. In contrast, the calcium gradient-dependent paracellular transport and calcium permeability were increased by PRL. Effects of PRL on Caco-2 monolayer were abolished by PI3K inhibitors (LY-294002 and wortmannin), but not by inhibitors of MEK (U-0126) or JAK2 (AG-490). To investigate whether the PRL-enhanced paracellular transport was linked to changes in the epithelial charge selectivity, the permeability ratio of sodium and chloride ([P.sub.Na]/[P.sub.Cl]) was determined. We found that PRL elevated the [P.sub.Na]/[P.sub.Cl] in both epithelia, and the effects were blocked by P13K inhibitors. In conclusion, PRL directly and rapidly stimulated the active and passive calcium transport in the rat duodenum and Caco-2 monolayer via the nongenomic P13K-signaling pathway. This PRL-enhanced paracellular calcium transport could have resulted from altered charge selectivity. charge selectivity; dilution potential; paracellular transport; prolactin receptor; tight junction; transcellular transport doi:10.1152/ajpendo.00142.2007

Abstract: 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 [...]

High physiological prolactin induced positive calcium balance by stimulating intestinal calcium absorption, reducing renal calcium excretion, and increasing bone calcium deposition in female rats. Although prolactin-induced increase in trabecular bone calcium deposition was absent after ovariectomy, its effects on cortical bones were still controversial. The present investigation, therefore, aimed to study the effect of in vivo long-term high physiological prolactin induced by either anterior pituitary (AP) transplantation or 2.5 mg/kg prolactin injection on cortical bones in ovariectomized rats. Since the presence of prolactin receptors (PRLR) in different bones of normal adult rats has not been reported, we first determined mRNA expression of both short- and long-form PRLRs at the cortical sites (tibia and femur) and trabecular sites (calvaria and vertebrae) by using the RT-PCR. Our results showed the mRNA expression of both PRLR isoforms with predominant long form at all sites. However, high prolactin levels induced by AP transplantation in normal rats did not have any effect on the femoral bone mineral density or bone mineral content. By using [sup.45]Ca kinetic study, 2.5 mg/kg prolactin did not alter bone formation, bone resorption, calcium deposition, and total calcium content in tibia and femur of adult ovariectomized rats. AP transplantation also had no effect on the cortical total calcium content in adult ovariectomized rats. Because previous work showed that the effects of prolactin were age dependent and could be modulated by high-calcium diet, interactions between prolactin and these two parameters were investigated. The results demonstrated that 2.0% wt/wt high-calcium diet significantly increased the tibial total calcium content in 9-wk-old young AP-grafted ovariectomized rats but decreased the tibial total calcium content in 22-wk-old adult rats. As for the vertebrae, the total calcium contents in both young and adult rats were not changed by high-calcium diet. The present results thus indicated that the adult cortical bones were potentially direct targets of prolactin. Moreover, the effects of high physiological prolactin on cortical bones were age dependent and were observed only under the modulation of high-calcium diet condition. bone mineral density; 45-calcium kinetic study; immunohistochemistry; pituitary transplantation; prolactin receptor

Abstract: Although an increase in trabecular-bone calcium deposition has been shown to be regulated by prolactin during lactation, the physiological significance of prolactin in bone calcium metabolism in nonlactating rats [...]