The effects of all-trans retinoic acid on glycosaminoglycan (GAG) accumulation were determined in cultured primary human skin fibroblasts. Confluent cultures treated with retinoic acid accumulated less [3H]GAG than those without the compound, an effect with an apparent threshold of 10 nM which was dose dependent in the concentration range tested (0-10 microM). At 10 microM, the inhibition was 54%. Greater than 80% of the labeled macromolecular material was streptomyces hyaluronidase digestible in cultures labeled with [3H]acetate. The incorporation of H2[35S]O4 into chondroitin sulfate and dermatan sulfate was unaffected, as was total protein synthesis. Retinol also inhibited accumulation of [3H]GAG, but was far less potent. T3 and dexamethasone can inhibit [3H]hyaluronate synthesis. When retinoic acid was added to cultures treated with either of these hormones at concentrations that maximally inhibit [3H] GAG accumulation, there was a further decrease in the rate of macromolecular accumulation. The retinoic acid effect evolved over 24-48 h after addition to the culture medium. A pulse-chase study failed to demonstrate any effect on [3H]GAG degradation.