Many men with erectile dysfunction have been successfully treated with intracavernosal injection of prostaglandin E(1) (PGE(1)) but this treatment is ineffective in 30 to 40% of patients. The goals of this study were to characterize PGE(1)-induced relaxation of isolated human penile smooth muscle (penile arteries and trabecular strips), correlating this in vitro response with the clinical response to this drug, and to evaluate the effects of the combination of PGE(1) with S-nitrosoglutathione (SNO-Glu) on relaxation of isolated human penile smooth muscle. Large variability in the EC(50) and maximal relaxation induced by PGE(1) was observed between tissues of different patients. Patients with poor clinical response to intracavernosal alprostadil (PGE(1)) had significantly larger EC(50) values and smaller maximal relaxation compared with patients with partial or complete clinical response to this drug. SNO-Glu consistently produced complete or near complete relaxation of human corpus cavernosum strips and penile arteries, even when the tissue responded poorly to PGE(1). In trabecular strips, the combination of PGE(1) and SNO-Glu in a 1:100 ratio demonstrated a synergistic relaxation effect. The combination of PGE(1) and SNO-Glu simultaneously increased the levels of both cAMP and cGMP in human corpus cavernosum tissue. Our results suggest that the clinical effectiveness of intracavernosal administration of PGE(1) is related to the variability of the relaxation responses of human trabecular tissue and penile arteries to this drug. The synergistic interaction of PGE(1) and SNO-Glu makes this combination an effective method to cause penile smooth muscle relaxation, a necessary step to initiate and maintain penile erection.