The neurotoxic effect of acute ethanol treatment (AET) may lead to an alteration in the regulation of calcium (Ca2+) homeostasis in hippocampal neurons. Ca2+ homeostasis could be affected by AET when neurons are at rest or after depolarizing activity during synaptic transmission. It has been shown that nerve growth factor (NGF) can ameloriate some types of neurotoxicity by stabilizing Ca2+ homeostasis. Previously, we observed that ethanol (EtOH) changed unstimulated (basal) and potassium (K+)-stimulated intracellular calcium ([Ca2+]i) in embryonic septohippocampal neurons (Webb et al., Brain Res. 729:176-189, 1996). The purpose of the present study is to determine the effects of NGF and EtOH on neuronal Ca2+ homeostasis in cultured embryonic hippocampal neurons. The hypotheses tested were the following: EtOH alters Ca2+ homeostasis in hippocampal neurons; NGF modulates Ca2+ homeostasis in hippocampal neurons; and NGF treatment alters the effect of EtOH on [Ca2+]i in hippocampal neurons. Our results indicated that hippocampal neuronal cultures treated with EtOH had lower basal [Ca2+]i than untreated neurons. EtOH decreased K+-stimulated (30 mM KCI) changes in [Ca2+]i in a dose-dependent manner. During K+ stimulation, 20 ng/ml of NGF slowed and reduced the increase in [Ca2+]i. Hippocampal neurons treated with NGF increased or did not change basal [Ca2+]i and did not change or increase K+-stimulated [Ca2+]i in response to EtOH. These responses were dose-related and indicated that NGF could alter the response of hippocampal neurons to EtOH. In conclusion, AET results in the alteration of Ca2+ homeostasis in unstimulated and depolarized cultured embryonic hippocampal neurons. NGF and EtOH independently and collectively affected the regulation of Ca2+ homeostasis in this neuronal population. Changes in [Ca2+]i can disrupt normal cellular function and contribute to cell death. Therefore, alteration of Ca2+ homeostasis may be an underlying mechanism involved in EtOH toxicity. NGF may ameliorate the toxic effects of EtOH by regulating Ca2+ homeostasis.