An increasing number of infants and children undergo surgery and are exposed to anesthesia as a part of medical care each year. Isoflurane is a commonly used anesthetic in the pediatric population. However, previous studies have reported widespread isoflurane-induced neuroapoptosis and cognitive impairments in neonatal animal models, raising concerns over the administration of isoflurane in the pediatric population. The current study investigated the effects of rutin, a flavonoid, on isoflurane-induced neuroapoptosis in a neonatal rodent model. Groups of neonatal rat pups were administered rutin at doses of 10, 20 or 40 mg/kg body weight from postnatal day 1 (P1) to P15. On P7, pups were exposed to 0.75% isoflurane for 6 h. Rat pups in the control groups did not receive rutin, and did not receive anesthesia in one group. Neuroapoptosis following isoflurane exposure was determined by TUNEL assay. The expression levels of cleaved caspase-3, apoptotic pathway proteins [Bcl2-associated agonist of cell death (Bad), phospho-Bad, Bax, B-cell lymphoma 2 (Bcl-2) and Bcl-xL and mitogen-activated protein kinases (MAPK)] signalling pathway proteins [c-Jun N-terminal kinase (JNK), phospho-JNK, extracellular-signal-regulated kinase 1/2 (ERK1/2), phosphoERK1/2, p38, phospho-p38 and phospho-c-Jun], were determined by western blot analysis. The Morris water maze test was used to assess the learning and memory of pups on P30 and P31. The present study found that rutin at the tested doses of 10, 20 and 40 mg significantly reduced (P<0.05) the isoflurane-induced elevation in apoptotic cell count. The expression levels of caspase-3, Bad, Bax and MAPK proteins, which were increased following isoflurane treatment, were rescued by rutin treatment. Furthermore, rutin prevented the increase in Bcl-xL, Bcl-2 and phospho-Bad expression following isoflurane treatment, and enhanced the memory of the rats. Rutin provided neuroprotection against isoflurane-induced neuronal apoptosis and improved the learning and memory of rats by effectively regulating the expression levels of proteins in the MAPK pathway.