Alternate bearing is a significant economic problem that affects fruit producers, consumers and the economy of a country. Concentrations of cytokinin (CK)-like compounds in different tissues of pistachio trees (Pistacia vera 'Uzun') were measured to elucidate the relationship between endogenous CKs and flower bud abscission (alternate bearing). Identification and quantification of CK-like compounds of the Z, DHZ and iP types from different physiological stages and tissues were performed using high-performance liquid chromatography-electrospray ionisation-tandem mass spectrometry (HPLC-ESI-MS/MS) techniques. Five to seven different CK-like compounds were detected, respectively. Isopentenyl adenine riboside (iPR) and trans-zeatin-O-glucoside (t-ZOG) were the main isoprenoid CKs, as the dominant conjugates in all the analysed organs of pistachio trees. The leaves and shoots of 'On'-year trees had significantly higher CK contents than those of 'Off'-year trees on all the sampling dates. Interestingly, the relatively high CK activity was recorded before flower bud abscission in the fractions corresponding to iPR and t-ZOG, while this activity dropped considerably during the flower bud abscission period (55 DAFB). Consequently, the strong negative correlation between CK concentration and bud abscission could be the first indication of the role of CK-like compounds in this unique phenomenon in the 'Uzun' pistachio cultivar. It was concluded that CKs could have an important role in the alternate bearing in pistachio.