It has been recently shown that the fast non-genomic responses of 1,25(OH)(2)-vitamin D-3 [1,25(OH)(2)D-3] in skeletal muscle cells involve tyrosine phosphorylation of MAP kinase (ERK1/2), c-Src kinase and the oncoprotein c-myc. In the present work, blockade of vitamin D receptor (VDR) expression (greater than or equal to80%) by preincubation of chick embryonic muscle cells with three different antisense oligonucleotides against the VDR mRNA (AS-VDR ODNs) significantly reduced (-94%) 1,25(OH)(2)D-3 stimulation of c-myc tyrosine phosphorylation and inhibited c-Src tyrosine dephosphorylation implying lack of c-Src activation by the hormone. Co-immunoprecipitation experiments revealed that 1,25(OH)(2)D-3 induces the formation of complexes between c-Src and c-myc, in agreement with the above results and previous studies showing hormone-dependent association between c-Src and tyrosine phosphorylated VDR and c-Src mediated c-myc tyrosine phosphorylation. MAPK tyrosine phosphorylation by 1,25(OH)(2)D-3 was affected to a lesser extent (-35%) by transfection with AS-VDR ODNs implying that both VDR-dependent and VDR-independent signalling mediate hormone stimulation of MAPK. These are the first results providing direct evidence on the participation of the VDR in non-genomic 1,25(OH)(2)D-3 signal transduction. Activation of tyrosine phosphorylation cascades through this mechanism may contribute to hormone regulation of muscle growth.