Glutathione S-transferases (GSTs) are a family of detoxification isoenzymes present in different tissues including the testis and that conjugate many toxic substrates to glutathione. Among these substrates are carcinogens, mutagens and products of oxidative processes. In the present report we show that GSTα is expressed in somatic testicular Leydig cells and Sertoli cells. GSTα expression in Sertoli cells is under the hormonal control of FSH, testosterone, and estradiol. In Leydig cells, immunoreactive GSTα was present at the neonatal, pubertal, and adult periods. In Sertoli cells, GSTα was predominant in pubertal and adult testes (but not in neonatal testes), suggesting that its expression is controlled by gonadotropins. The regulatory action and the mechanisms of action of FSH and testosterone on GSTα mRNA and protein levels were studied by using a model of primary cultures of porcine testicular Sertoli cells. FSH increased GSTα mRNA levels in a dose-dependent manner (ED₅₀ = 18.5 nm/ml) with a maximal effect observed after 48 h of exposure (a 3-fold increase; P < 0.001). In addition, FSH increased GSTα protein, which was detected as a doublet of 28 kDa. Treatment with testosterone enhanced GSTα mRNA levels in a dose-dependent (ED₅₀ = 1.4 ng/ml) and time-dependent manner with a maximal effect delayed at 8 h of exposure (a 2-fold increase; P < 0.001). Similarly, Sertoli cell treatment with testosterone metabolites, dihydrotestosterone (DHT) and estradiol, led to an increase in GSTα mRNA levels. Because stimulatory effects of FSH and androgens were also observed on GSTα protein, we therefore had to determine whether the different hormones were affecting GSTα gene transcriptional activity, or GSTα mRNA stability, or both. FSH and 8-Br-cAMP (but not testosterone) increased the stability of GSTα mRNA. The effects of FSH and testosterone on GSTα protein were additive, confirming that both hormones act through distinct mechanisms on the expression of the enzyme. Taken together, the present observations indicate that Sertoli cell GSTα is targeted by FSH, testosterone, and its metabolites, and they reinforce the concept that Sertoli cells exert a protective role and are under endocrine control to ward against toxic agents in the context of Sertoli-germ cell interactions during spermatogenesis.