The role of microtubules in modulating cardiomyocyte beta-adrenergic response was investigated in rats with cardiac hypertrophy. Male Sprague-Dawley rats underwent stenosis of the abdominal aorta (hypertensive, HT) or sham operation (normotensive, NT). Echocardiography and isolated left ventricular cardiomyocyte dimensions demonstrated cardiac hypertrophy in the HT rats after 30 wk. Cardiomyocyte microtubule fraction was assayed by high-speed centrifugation and Western blot. In contrast to previous reports of increased microtubules after acute pressure overload, microtubule fraction for HT was significantly lower than that for NT. Cardiomyocytes were exposed to either 1 microM colchicine, 10 microM taxol, or equivalent volume of vehicle. Colchicine decreased microtubules, and taxol increased microtubules in both groups. Cardiomyocyte cytosolic calcium ([Ca2+]c) and shortening/relaxation dynamics were assessed during exposure to increasing isoproterenol concentrations. The beta-adrenergic response for these variables in the HT group was blunted compared with NT. However, increased microtubule assembly by taxol partially recovered the normal beta-adrenergic response for time to peak [Ca2+]c, time to peak shortening, and mechanical relaxation variables. Microtubule assembly may play a significant role in determining cardiomyocyte beta-adrenergic response in chronic cardiac hypertrophy.