Cortical spreading depression (CSD) is an important disease model for migraine and cerebral ischemia. The spatio-temporal characteristics of the intrinsic optical signals (IOS) at 540 nm and 660 nm during CSD were investigated by high resolution optical reflectance imaging through thinned skull in rats. The possible physiological mechanisms underlying the IOS were discussed. The CSD were induced by pinprick in 16 α-chloralose/urethane anesthetized Sprague-Dawley rats. A four-phasic IOS response of decreased ［amplitude(－2.1±1.2)%, duration(16.2±3.8)s］, increased ［amplitude(2.9±1.6)%, duration (13.8±2.2)s］, decreased ［amplitude(－14.2±4.5)%, duration(40.6±8.4)s］ and increased (146.2±40.3)s optical reflectance was observed at pial arteries and parenchyma sites in all experimental animals at 540 nm. The IOS spreads peripherally from the site of CSD induction at speeds of (3.7±0.4) mm/min, companying a dramatic dilation (69.2±26.1)% of the pial arteries. At 660 nm, a three-phasic IOS response of increased ［amplitude(3.8±0.6)%, duration(17.9±5.1)s］, decrease ［amplitude(－3.0±1.7)%, duration(43.3±6.4)s］ and increased optical reflectance was recorded at parenchyma sites.