The effect of heparin on the amount of membrane-associated heparan sulfate proteoglycan (HSPG) was examined biochemically in balloon-injured rat carotid arteries. Heparin (Choay 1772) was administered at the rate of 1 mg/kg/h i.v. by an osmotic infusion pump for 10 days. Arterial tissues were incubated ex vivo with Na<sub>2</sub><sup>35</sup>SO<sub>4</sub> and <sup>3</sup>H-glucosamine for 6 or 24 h. The arteries from heparin-treated rats were also incubated with heparin (l00 µg/ml) during the radiolabeling. Membrane-associated proteoglycans were extracted with 4 M guanidine-HCl and isolated by a series of chromatographic steps using Sephadex G-50, DEAE-Sephacel and Octyl-Sepharose CL-4B. Membrane-associated proteoglycans were eluted with 0.8% Triton X-100 from Octyl-Sepharose columns. Heparin treatment significantly increased membrane-associated HSPG by 64% in the arteries incubated for 24 h, while heparin hardly increased the HSPG in 6 h incubated arteries. Since the 24-hour data seem to reflect a combination of biosynthesis and degradation of proteoglycans while the 6-hour data primarily reflect biosynthesis, these results suggest that inhibition of degradation of membrane-associated HSPG is involved in the mechanism of heparin action. This speculation is supported by the observation that membrane-associated HSPG was elevated in balloon-injured arteries by treatment with protease inhibitors.