Biomedical subjects

P Lembessis

Publications and source records attributed to P Lembessis.

3 recordsLinked to original sources

PubMed1992

Differences in Ca(2+)-mediation of hypotonic and Na(+)-nutrient regulatory volume decrease in suspensions of jejunal enterocytes.

We determined differences in the Ca2+ signalling of K+ and Cl- conductances required for Regulatory Volume Decrease (RVD) in jejunal villus enterocytes passively swollen (0.5 or 0.95.isotonic) compared with swelling because of the absorption of D-glucose (D-Glc) or L-Alanine (L-Ala). Cell volume was measured using electronic cell sizing. In nominally Ca(2+)-free medium containing EGTA (100 microM) RVD after 0.5 or 0.95.isotonic challenge was prevented. L-Ala swelling and subsequent RVD was influenced in Ca(2+)-free medium. Villus cells were incubated with 10 microM of the acetomethoxy derivative of 1,2.bis (2-aminophenoxy) ethane N,N,N1,N1 tetracetic acid (BAPTA-AM) and RVD after 0.5.isotonic swelling or L-Ala swelling was prevented. Niguldipine (0.1 microM), nifedipine (5 microM), diltiazem (100 microM), Ni2+, and Co2+ (1 mM) all prevented hypotonic RVD but had no effect on RVD after L-Ala addition. Charybdotoxin (25 nM) a potent inhibitor of Ca(2+)-activated K+ channels, had no effect on hypotonic RVD but prevented RVD of villus cells swollen by D-Glc. We used the calmodulin antagonists, naphthalene sulfonamide derivatives W-7 and W-13, to assess calmodulin activation of K+ and Cl- conductance in these two models. L-Ala swelling and subsequent RVD was not influenced by 25 microM W-7; hypotonic RVD was prevented by 25 microM W-7 or 100 microM W-13. The W-13 inhibition of RVD was by-passed with 0.5 microM gramicidin. Our data show that hypotonic RVD requires extracellular Ca2+ and that the K+ conductance activated is not charybdotoxin sensitive but requires calmodulin.(ABSTRACT TRUNCATED AT 250 WORDS)

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PubMed1992

Effect of protein kinase C inhibitors on Cl- conductance required for volume regulation after L-alanine cotransport.

We used electronic cell sizing and Cl- efflux measurements in guinea pig jejunal enterocytes to study activation of Cl- conductance under two experimental conditions, regulatory volume decrease (RVD) after passive hypotonic swelling and volume regulation during Na(+)-alanine cotransport. RVD after a hypotonic (0.5 x isotonic) challenge was not affected by the protein kinase C (PKC) inhibitor 100 microM 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7). Volume decrease after cell swelling in response to L-Ala (25 mM) was prevented by H-7 (P less than 0.05) or the more potent PKC inhibitor 10 nM staurosporine (P less than 0.001). L-Ala stimulated biphasic 36Cl efflux, a rapid efflux over 60 s which was inhibited by H-7 (P less than 0.01) and the Cl(-)-channel blocker anthracene-9-carboxylic acid (9-AC) (P less than 0.005). In contrast, after hypotonic dilution the rate of 36Cl efflux increased (P less than 0.005); H-7 had no effect but 9-AC inhibited the increase (P less than 0.01). Gramicidin (0.5 microM) added to cells maximally swollen by L-Ala in Cl(-)-containing medium caused 2 degree swelling (P less than 0.001), but 10 nM staurosporine reduced this 2 degree swelling (P less than 0.001). Addition of phorbol ester or synthetic diacylglycerol to villus cells under isotonic conditions, after gramicidin addition, caused cell swelling (P less than 0.005) that was inhibited by staurosporine (P less than 0.05). We concluded that PKC does not activate Cl- conductance for hypotonic RVD but that Na(+)-nutrient cotransport is a physiological stimulus for PKC to activate Cl- conductance necessary for volume regulation.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1992

Effect of osmotic swelling on K+ conductance in jejunal crypt epithelial cells.

To further elucidate differences in ion transport properties between jejunal crypt and villus cells, we compared the responses of purified cell suspensions to hypotonic stress using electronic cell sizing to evaluate volume changes and 86Rb and 36Cl efflux. After hypotonic swelling, villus enterocytes undergo a regulatory volume decrease (RVD) due to the loss of K+ and Cl- through volume-activated conductances. After 0.6x isotonic challenge in Na(+)-free medium, crypt cells exhibited only partial RVD, with t1/2 congruent to 15 min. The addition of a cation ionophore, gramicidin (0.25 microM), to hypotonically swollen crypt cells caused an accelerated RVD, which was complete with t1/2 congruent to 5 min. Crypt epithelial cells showed no volume-activated 86Rb efflux, but villus enterocytes had an increased rate of 86Rb efflux after hypotonic dilution (P less than 0.001). Gramicidin added to hypotonically diluted crypt cells greatly increased the rate of 86Rb efflux compared with controls. Both villus (30 s; P less than 0.005) and crypt (2 min; P less than 0.001) cells exhibited volume-activated 36Cl efflux in absence of gramicidin. Cl- channel blockers anthracene-9-carboxylate (9-AC, 300 microM) and indanyloxyacetic acid (IAA-94, 100 microM) prevented crypt RVD (P less than 0.001) in the presence of gramicidin. Ouabain (P less than 0.001) or K(+)-free Na(+)-containing medium, but not Ba2+ (5 mM) or quinine (100 microM), prevented crypt partial RVD. We conclude that crypt cells lack volume-activated K+ conductance. The RVD exhibited by crypt cells, although partial, was due to Cl- loss through a volume-activated Cl- conductance and Na+ loss via Na(+)-K(+)-ATPase.

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