Background:Transzonal projections (TZPs) constitute a structural basis for the communication between the oocyte and its surrounding cumulus cells (CCs), which play critical roles in promoting the oocyte maturation. Previously we found that heat stress (HS) causes loss of TZPs in porcine cumulus-oocyte complexes (COCs) with decreased density of filamentous actin (F-actin). However, the time-course responses of F-actin and its monomeric actins (β-actin andγ-actin) during the in vitro maturation of oocytes remain unclear. Results:In this study, excised porcine ovaries were exposed to HS at 41.5 °C for 1 h before COCs were isolated and matured in vitro for 44 h. HS significantly reduced oocyte quality, characterized by impaired cumulus expansion, delayed meiotic resumption and lower survival rate and polar body extrusion rate, as well as decreased expression of mitochondrial DNA-encoded genes and elevated mitochondrial reactive oxygen species concentration. Expression ofβ-actin andγ-actin in CCs increased gradually with oocytes maturation, which was significantly reduced in HS group, especially at 24 h and/or 44 h of in vitro maturation. By contrast, the number of TZPs and the fluorescence intensity of F-actin in zona pellucida decreased gradually during oocytes maturation, which were significantly reduced by HS at 24 h of in vitro maturation. Moreover, colocalization analyses revealed bothβ-actin andγ-actin contribute to the F-actin formation in porcine TZPs, and the colocalization of F-actin with GJ protein connexin 45 was significantly reduced in heat-exposed COCs. Conclusions:The results indicate that the suppression of actin expressions in CCs, which may lead to the F-actin unstabilization in TZPs, will subsequently contribute to the compromised quality of oocytes under HS.