Single-channel and whole-cell recordings of mitogen-regulated inward currents in human cloned helper T lymphocytes

Abstract

Cytoplasmic free Ca²⁺ ([Ca²⁺]_i) appears to be an important signal for DNA synthesis in early stages of lymphocyte activation. In spite of many experimental studies which employ fluorescent Ca²⁺ indicator dye to demonstrate an early increase of [Ca²⁺]_i in T-lymphocytes after stimulation with lectins^1–3, specific antigens^4,5, and monoclonal antibodies to T-lymphocyte receptors^6–8, the mechanism responsible for the rise of [Ca²⁺]_i is unknown. We have used the extracellular patch clamp technique⁹ to investigate this mechanism. Unitary inward currents, mediated by Ca²⁺ or Ba²⁺, were recorded in the membrane of T-lymphocytes. The inward current channel was characterized by a conductance of 7 pS and extrapolated reversal potential (E_rev) 110mV positive to resting potential (V_r). While gating kinetic parameters were not affected by membrane potential changes, the probability of channel opening markedly increased upon activation of the T-lymphocyte by the mitogenic lectin, phytohaemagglutinin (PHA). PHA also evoked a cadmium-sensitive, inward Ba²⁺ current on whole-cell clamp. We suggest that this mitogen-regulated channel introduces Ca²⁺ into the cytoplasm upon activation and represents a new class of voltage-independent Ca²⁺ channels.