Grayanotoxins (GTXs) are natural products and are mostly found in plants of the Ericaceae family, especially in the Rhododendron. With their ability to bind to voltage-gated sodium channels, they keep these channels constantly active and cause tissue damage. However, despite this feature, the use of Rhododendron leaves or its secondary products as an alternative product is especially common in Turkey. This study aims to evaluate the possible dose-related effects of GTX-III in ischemia-induced in vitro cell models. Within the scope of the study, an ischemia model was established in two different cell lines (H9c2 and Cos-7) and treated with various concentrations of GTX-III. In this context, cell viability, cytotoxicity, apoptosis and necrosis were examined. In the results of MTT, a significant decrease (+p < 0.05) in cell viability was observed in all GTX-III concentrations in H9c2 cells compared to the control, while a significant difference (+p < 0.05) was observed in Cos-7 cells, especially at the 24th hour. LDH cytotoxicity was increased in a dose-dependent manner in both cell models. It was concluded that GTX-III caused apoptosis, and reduced cell viability in ischemia models; however, promoted cell proliferation in healthy cells. Based on the literature review, this study is the first to document the cytotoxic properties and apoptotic potential of GTX-III in an in vitro cell culture ischemia model. Our findings support the usage of GTX-III, however it should be remembered that the dose needs to be verified before being used medically.