Our previous study demonstrated significant difference in the basal plasma cortisol levels between Erhualian (EHL) and Pietrain (PIE) pigs, implicating fundamental breed difference in adrenocortical function. The objectives of the present study were therefore to characterize the expression pattern of proteins involved in adrenal ACTH signaling and, including melanocortin type 2 receptor (MC2R), cAMP response element binding protein (CREB) and phosphorylated CREB (pCREB), steroidogenic acute regulatory protein (StAR), as well as that of the key enzymes involved in steroidogenesis in EHL and PIE pigs, in association with the plasma corticotrophin (ACTH) and cortisol levels. The plasma concentrations of the substrates for adrenal steroidogenesis, cholesterol and low-density lipoprotein (LDL) cholesterol, did not differ between breeds. Plasma concentration of ACTH and the adrenal contents of MC2R mRNA and protein were similar in two breeds of pigs, whereas the basal plasma concentrations of cortisol in EHL pigs were 1.5 folds higher than that in PIE pigs. The higher basal plasma cortisol levels in EHL pigs were found to be accompanied with the higher expression of ACTH post-receptor signaling components, cAMP, pCREB and StAR, as well as the higher expression of cholesterol side-chain cleavage cytochrome P450 (P450scc), 17alpha-hydroxylase cytochrome P450 (P450(17alpha)), 21-hydroxylase cytochrome P450 (P450c21) and 11beta-hydroxylase cytochrome P450 (P450(11beta)). These results indicated that the enhanced cAMP/PKA/pCREB-signaling system and augmented expression of StAR and steroidogenic enzymes are major attributes to the higher basal plasma cortisol concentrations in pigs.