The majority of left ventricular (LV) inflow volumes in hypertrophic cardiomyopathy (HCM) depend on atrial contraction because of impaired LV relaxation. If HCM is complicated by atrial fibrillation (AF), heart failure can develop because of the loss of atrial contraction. The purpose of this study was to determine the relationship between the development of AF and myocardial fibrosis or intramyocardial small artery (IMSA) stenosis in autopsied hearts with HCM. Studies were performed in five HCM hearts with AF (AF group) and five HCM hearts without AF (non-AF group). LV specimens were divided into the inner (IT), middle (MT), and outer (OT) thirds. We selected at random 120 fields and 20 IMSAs from each layer and assessed them quantitatively using an image analyzer. We determined the extent of fibrosis (%F) and the degree of stenosis of each IMSA (%L). The %F in the AF group was greater than in the non-AF group (P<.01). In the AF group, the %F of the IT was greater than in the MT and the OT (P<.01). In the non-AF group, the %F of the IT was greater than in the MT (P<.05), and the %F of the MT was greater than in the OT (P<.01). The %L was similar in the AF and non-AF groups. In both groups, the %L of the IT was lower than in the MT (P<.01), which was lower than that of the OT (P<.05). LV fibrosis is more severe in patients with HCM and AF than in those without AF. Therefore, myocardial fibrosis might impair LV relaxation, resulting in hemodynamic intolerance to AF.