Basolateral membranes isolated from rat jejunal enterocytes and enriched 14 times were found present as unsealed and sealed right side out (RSO) or inside out (IO) vesicles in the ratio 2:2:1. ...Entrance of 1 mM Na (JNa) into basolateral membrane vesicles was measured in the presence and in the absence of 5 mM ATP by a rapid filtration technique, under different experimental conditions. JNa across the basolateral membrane is cis-inhibited by K, transstimulated by K and further stimulated by ATP (activation of the Na pump). Both K- and ATP-positive effects are also obtained by other cations: the relative stimulation sequence is K greater than Rb = NH4 greater than Cs. The ATP effect can be suppressed by vanadate and strophanthidin; moreover the K effect on JNa is not influenced by blocking the mechanism of the Na pump with ouabain, strophanthidin and adenosine 5'-beta,gamma-imidotriphosphate. The transmembrane potential does not influence the K-stimulated JNa. In addition to the Na pump this study demonstrates the existence of an exchange mechanism between Na and K, which seems to be different from the Na pump. There appears to be no Na-K cotransport.
Basolateral membranes from rat jejunal enterocytes have been obtained by self-orienting Percoll-gradient centrifugation. Bicarbonate and L-glucose uptake into osmo-tically active basolateral membrane ...vesicles has been studied by a rapid filtration technique. In closed vessels and at pH 8.2 the uptake kinetics of both
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Cbicarbonate and L
3
Hglucose have been followed for 30 min at 18 °C. Bicarbonate uptake seems to be fast and in efflux experiments SITS and DIDS effect is negligible. This work demonstrates that it is possible to determine bicarbonate flux across basolateral membrane vesicles at pH and temperature values close to usual experimental conditions.
4-Acetamido-4'-isothiocyano-2,2'-disulfonic stilbene (SITS) and 4,4'-diisothiocyano-2,2'-disulfonic stilbene (DIDS) effects have been tested both in the basolateral membranes (BLMs) of jejunum ...enterocytes and in the same intestinal tract, everted and incubated "in vitro". Total and (Na,K)-ATPase activities of BLMs are inhibited in a similar way by the two disulfonic stilbenes as well as the fluid transintestinal transport in the everted intestine; on the contrary cell Na and K are unaffected. SITS and DIDS inhibition of (Na,K)-ATPase seems to take place at the cytoplasmic side of the BLM.
Evidence was previously given that the mechanisms involved in bicarbonate and lactate movements across rat jejunal enterocyte are modulated by PKC and Ca2+/CaM. Aim of this study was to investigate ...the possible role of PKA on bicarbonate and lactate transports.
Enzymatic assays in isolated plasma membranes were performed. Moreover membrane vesicles, transiently opened and resealed, were treated with a phosphorylating solution (leading to PKA activation) and were used after that to perform uptake studies.
Enzymatic assays give evidence for the presence of PKA in plasma membranes from rat jejunum. Uptake experiments suggest that PKA stimulates the two systems that accomplish basolateral HCO3- efflux from the enterocyte, namely Cl-/ HCO3- exchanger and HCO3- conductance, without affecting HCO3- influx from the lumen mediated by Na+/H+ exchanger activity. Moreover basolateral H+-lactate symporter is stimulated by PKA, as well as the brush border isoform of Na+-glucose cotransporter SGLT1.
PKA activation evokes individual responses that could be coordinated through cellular metabolism.
