SODIUM ION-DEPENDENT (N-METHYLAMINO)-ALPHA-ISOBUTRYIC ACID UPTAKE BY EMBRYONIC CHICK-CELLS EXPOSED TO ETHANOL IN OVO - RESPONSE TO THE STIMULATION DOWNREGULATION OF PROTEIN-KINASES/

Amino acid uptake, critical for embryonic development, was studied in cultured chick cells isolated from 5-day-old chick embryos that had re ceived treatment in ovo with either vehicle (sterile chick Ringer's so lution) or Vehicle plus ethanol (1.5 g/kg). Upon being placed in cultu re, the cells were grown in the absence of ethanol per se. The uptake of (N-methylamino)-alpha-isobutryic acid (AIB) was used to estimate am ino acid uptake via system A, a Na+ dependent system that transports s hort-chain amino acids. The uptake of AIB in cultured chick embryo cel ls in response to the stimulation/ downregulation of three protein kin ases [tyrosine kinase, cyclic AMP-dependent kinase A (PKA) and protein kinase C (PKC)] was determined. Acute activation of PKC by treatment of the cells with a phorbol ester (TPA) inhibited Nac-dependent AIB up take. Conversely, treatment of the cells with TPA for 24 h, to downreg ulate PKC activity, significantly increased AIB uptake. The data sugge st that in these cells, PKC plays an important role in the regulation of amino acid uptake via system A. Activation of PKA via treatment of the cells with forskolin, an adenylyl cyclase activator, had little ef fect on AIB uptake. in cells from vehicle-treated embryos and only a s light depressing effect on uptake by cells from ethanol-treated embryo s. Insulin and insulin-like growth factor-1 (IGF-1) both stimulated AI B uptake, but the higher concentrations of insulin necessary to increa se uptake to a level comparable to that of IGF-1 stimulation suggested that insulin could be acting through the IGF-1 receptor. Thus, while AIB uptake by cells of in ovo ethanol-treated embryos was significantl y increased by treatment with either insulin or IGF-1, IGF1 appeared t o be the more physiologically important compound in the chick embryo. Overall, there was a consistent trend for cells isolated from in ovo e thanol-treated embryos to have higher levels of AIB uptake relative to cells isolated from vehicle-treated embryos, regardless of the in vit ro treatment.