To evaluate the comparative efficacy of Ovsynch and Heatsynch protocols with respect to ovarian response, estrus, pregnancy rate and progesterone concentrations, 29 cyclic buffalo heifers were assigned randomly to one of four treatments: GPG treatment (n= 8) received two GnRH doses, one seven days before and the other two days after a single dose of PGF2á; GP-EB1 treatment (n= 8) was given estradiol benzoate (EB) one day after PGF2á treatment instead of the second GnRH dose; GP-EB2 (n= 8) and GP-ECP (n= 5) treatments received EB or estradiol cypionate instead of second GnRH in GPG protocol, respectively. Ovarian ultrasonography and expression of estrus signs were monitored twice daily until the fifth day after the last hormone dose. Our findings indicated a more obvious estrus response to estradiol than the second GnRH dose. The longer estrus duration observed in response to estradiol (32 to 34 h) could provide more time for estrus detection and insemination in these animals.