Electrogenic Na⁺-dependentl-alanine transport in the lizard duodenum. Involvement of systems A and ASC

Abstract

L-Alanine transport across the isolated duodenal mucosa of the lizard Gallotia galloti has been studied in Ussing chambers under short-circuit conditions. Net L-alanine fluxes, transepithelial potential difference (PD), and short-circuit current (Isc) showed concentration-dependent relationships. Na(+)-dependent L-alanine transport was substantially inhibited by the analog alpha-methyl aminoisobutyric acid (MeAIB). Likewise, MeAIB fluxes were completely inhibited by L-alanine, indicating the presence of system A for neutral amino acid transport. System A transport activity was electrogenic and exhibited hyperbolic relationships for net MeAIB fluxes, PD, and Isc, which displayed similar apparent K(m) values. Na(+)-dependent L-alanine transport, but not MeAIB transport, was partially inhibited by L-serine and L-cysteine, indicating the participation of system ASC. This transport activity represents the major pathway for L-alanine absorption and seemed to operate in an electroneutral mode with a negligible contribution to the L-alanine-induced electrogenicity. It is concluded from the present study that the active Na(+)-dependent L-alanine transport across the isolated duodenal mucosa of Gallotia galloti results from the independent activity of systems A and ASC for neutral amino acid transport.

Regulation of L-alanine transport systems A and ASC by cyclic AMP and calcium in a reptilian duodenal model

“…As has been reported several times for this same preparation (Gómez et al, 1986;Lorenzo et al, 1989b), no differences were observed between the transmural potential difference (PD), short-circuit current (Isc) and tissue conductance (Gt) between stable open-circuit and short-circuit measurements. As can be seen in Table·1, under control conditions, transepithelial PD was approximately 2.0·mV and the mean Isc was 0.77·µequiv.·cm -2 ·h -1 , the serosal side of the tissue being electrically positive compared with the luminal side, which parallels our previous observations (Gómez et al, 1986;Medina et al, 2001). Both PD and Isc were significantly increased by forskolin, theophylline and db-cAMP, in agreement with most studies performed in different animal models, including lizard intestine (Bridges et al, 1983;Díaz and Lorenzo, 1991;Lorenzo et al, 1989a).…”

“…L-alanine transport The effects of different pharmacological agents affecting cAMP metabolism on unidirectional and net fluxes of L-alanine (1·mmol·l -1 ) across the isolated lizard duodenum have been determined under short-circuit conditions (Fig.·1). Using Na + -containing Ringer solutions, mean control net L-alanine transport (Jnet) was 14.81±1.05·nmol·cm -2 ·h -1 (significantly different from zero at P<0.01), which was in the range reported previously for this same preparation (Medina et al, 2001). Addition of forskolin (10·µmol·l -1 ), theophylline (3·mmol·l -1 ) or the membrane-permeable analogue dibutyryl cyclic AMP (db-cAMP; 500·µmol·l -1 ) significantly increased Jnet, these effects being due to a rise in the mucosal-to-serosal flux (Jms) without altering the flux in the opposite direction (Jsm).…”

“…This same procedure has been used to isolate the ASC transport pathway in different preparations, including lizard duodenum (Medina et al, 2001). Interestingly, under these conditions, both intracellular messengers affected L-alanine transport but in a completely opposite direction (Figs·4C, 5C).…”

“…We have previously demonstrated that, under short-circuit conditions, the overall L-alanine transport in the lizard duodenum could be completely explained by the simultaneous participation of two Na + -dependent transport systems, endowed with properties of systems ASC and A, plus one Na + -independent electrogenic carrier (Díaz et al, 2000;Medina et al, 2001). These three different transport systems could be individually dissected by means of a strategy based on the differential substrate affinities and sodium dependence.…”

“…This epithelium has been studied over the past two decades, and the different pathways for neutral amino acid transport have been recently identified and characterised physiologically (Díaz et al, 2000;Medina et al, 2001). …”