The intracellular mechanisms underlying T lymphocyte activation leading to demyelination in multiple sclerosis (MS) have not yet been clarified. We have recently reported that interferon (IFN)-gamma activates a novel trans-plasmalemma Ca2+ influx on T lymphocytes (mainly CD4+) from patients with MS which induces intracellular Ca2+ ([Ca2+]i) elevation. Since Ca2+ is an essential second messenger in regulating transcription of T lymphocyte activation genes, we have evaluated how [Ca2+]i elevation due to the activity of this particular influx affects T lymphocyte proliferative behaviour in 12 influx-positive relapsing-remitting MS (RR-MS) patients. Fourteen influx-negative RR-MS patients and 14 healthy donors were used as controls. In lymphocytes from healthy controls, a significant correlation (r = 0.62; P < 0.001) was found between [Ca2+]i levels and proliferation rate after phytohemagglutinin (PHA) stimulation. Sustained proliferation was induced in T lymphocytes by > or = 10 micrograms/ml of PHA, a dose leading to a [Ca2+]i increase of at least 45% over basal level. Similar [Ca2+]i elevations were obtained when > or = 10 micrograms/ml of PHA were used on cells from RR-MS patients. However, T lymphocytes from RR-MS patients, but not from healthy donors, proliferated also in response to 1 micrograms/ml of PHA, indicating a state of preactivation. Moreover, 1 microgram/ml of PHA used in combination with suboptimal doses of IFN-gamma (5 UI/ml) doubled the proliferation rate of influx-positive MS cells, but not influx-negative MS cells or cells from healthy donors compared to the values obtained using PHA alone (P < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)