Contents: The main scope of this manuscript is to analyse the dynamics of mitochondrial activity in boar sperm subjected to 'in vitro' capacitation (IVC) and subsequent progesterone-induced 'in vitro' acrosome reaction (IVAR). This was determined after analysis of the rhythm of O 2 consumption and concomitant changes in the mitochondria activity-specific JC-1 staining. Results showed that IVC, and especially IVAR, was concomitant with a peak in O 2 consumption (from 1.61±0.08nmol O 2/min/10 7 viable sperm at 0h of incubation to 2.62± 0.12nmol O 2/min/10 7 viable sperm after 5min of IVAR induction). These results were accompanied by parallel changes in the mean intensity of JC-1 staining. Based on JC-1, mitochondrial activation followed a nucleated pattern, with specific, activation starting points at the midpiece from which mitochondrial activation was spread. Moreover, four separate sperm subpopulations were detected following the JC-1 orange-red/green ratio, and the observed changes in the mean JC-1 staining during IVC and IVAR were related to concomitant changes in both the orange-red/green JC-1 ratio and the percentage of sperm included in each subpopulation. All of these results indicate that IVC and the first minutes of IVAR are accompanied by a progressive increase in mitochondrial activity, which reached a peak coincidental with the achievement of IVAR. Moreover, results suggest the presence of separate sperm subpopulations, which show a different mitochondrial sensitivity to IVC and IVAR. Finally, mitochondrial activation, at least under JC-1 staining, seems to originate in concrete nucleation points at the midpiece, thus suggesting thus a well-coordinated pattern in boar-sperm mitochondrial activity modulation. © 2010 Blackwell Verlag GmbH.