Calcium absorption by Ca_v1.3 induces terminal web myosin II phosphorylation and apical GLUT2 insertion in rat intestine

Abstract

Glucose absorption in rat jejunum involves Ca2+ -and PKC βII-dependent insertion of GLUT2 into the apical membrane. Ca 2+ -induced rearrangement of the enterocyte cytoskeleton is thought to enhance paracellular flow. We have therefore investigated the relationships between myosin II regulatory light chain phosphorylation (RLC 20 ), absorption of glucose, water and calcium, and mannitol clearance. ML-7, an inhibitor of myosin light chain kinase, diminished the phloretin-sensitive apical GLUT2 but not the phloretin-insensitive SGLT1 component of glucose absorption in rat jejunum perfused with 75 mM glucose. Western blotting and immunocytochemistry revealed marked decreases in RLC 20 phosphorylation in the terminal web and in the levels of apical GLUT2 and PKC βII, but not SGLT1. Perfusion with phloridzin or 75 mM mannitol, removal of luminal Ca 2+ , or inhibition of unidirectional 45 Ca 2+ absorption by nifedipine exerted similar effects. ML-7 had no effect on the absorption of 10 mM Ca 2+ , nor clearance of [14 C]-mannitol, which was less than 0.7% of the rate of glucose absorption. Water absorption did not correlate with 45 Ca 2+ absorption or mannitol clearance. We conclude that the Ca 2+ necessary for contraction of myosin II in the terminal web enters via an L-type channel, most likely Ca v 1.3, and is dependent on SGLT1. Moreover, terminal web RLC 20 phosphorylation is necessary for apical GLUT2 insertion. The data confirm that glucose absorption by paracellular flow is negligible, and show further that paracellular flow makes no more than a minimal contribution to jejunal Ca 2+ absorption at luminal concentrations prevailing after a meal.

CaV1.3 Channels and Intracellular Calcium Mediate Osmotic Stress-induced N-terminal c-Jun Kinase Activation and Disruption of Tight Junctions in Caco-2 Cell Monolayers

“…Ca V 1.3, is a major voltage-gated Ca 2ϩ channel in the intestinal epithelium. Ca V 1.3 is involved not only in Ca 2ϩ absorption (19), but also in Ca 2ϩ signaling leading to myosin light chain kinase activation (28). Our present study demonstrates that knockdown of Ca V 1.3 significantly reduces the osmotic stressinduced increase in Ca 2ϩ and abrogates osmotic stress-induced tight junction disruption and barrier dysfunction.…”

Transepithelial calcium transport in prolactin-exposed intestine-like Caco-2 monolayer after combinatorial knockdown of TRPV5, TRPV6 and Cav1.3

“…and glucose uptake through sodium-dependent glucose transporter 1 (SGLT1) at the apical membrane is coupled with Ca v 1.3 opening, which requires cell depolarization induced by SGLT1-mediated Na ? entry [8,37], it is also possible that PRL indirectly activates Ca v 1.3 by acting through SGLT1, known to be expressed in Caco-2 cells [38].…”

GLUT2 protein at the rat proximal tubule brush border membrane correlates with protein kinase C (PKC)-βl and plasma glucose concentration

“…It is now clear that sodium/glucose cotransporters (SGLTs) and GLUT2 work in conjunction to transport glucose across the brush border membrane (BBM) over a wide range of glucose concentrations. The proposed model for enterocyte glucose uptake is that a high level of luminal glucose leads to rapid insertion of GLUT2 into the BBM via a calcium-and PKC-βII-dependent mechanism [5][6][7][8]. There is less evidence to support a similar process occurring in the proximal tubule BBM, but because the basic glucose transport mechanisms are similar in both tissues, it seems likely that the same model may apply.…”