The M-type potassium current (I_M) plays a dominant role in regulating membrane excitability and is modulated by many neurotransmitters. However, except in the case of bradykinin, the signal transduction pathways involved in M-channel modulation have not been fully elucidated. The channels underlying I_M are produced by the coassembly of KCNQ2 and KCNQ3 channel subunits and can be expressed in heterologous systems where they can be modulated by several neurotransmitter receptors including histamine H₁ receptors. In HEK293T cells, histamine acting via transiently expressed H₁R produced a strong inhibition of recombinant M-channels but had no overt effects on the voltage dependence or voltage range of I_M activation. In addition, the modulation of I_M by histamine was not voltage-sensitive, whereas channel gating, particularly deactivation, was accelerated by histamine. Nonhydrolysable guanine nucleotide analogues (GDP--S and GTP--S) and pertussis toxin (PTX) treatment demonstrated the involvement of a PTX-insensitive G-protein in the signal transduction pathway mediating histamine-induced I_M modulation. Abrogation of the histamine-induced modulation of I_M by expression of a C-terminal construct of phospholipase C (PLC-1-ct), which buffers activated G_q/11 subunits, implicates this G protein subunit in the modulatory pathway. On the other hand, abrogation of the histamine-induced modulation of I_M by expression of two constructs which buffer free subunits, transducin (Gt) and a C-terminal construct of a G protein receptor kinase (MAS-GRK2-ct), implicates dimers in the modulatory pathway. These findings demonstrate that histamine modulates recombinant M-channels in HEK293T cells via a PTX-insensitive G-protein, probably G_q/11, in a similar manner to a number of other G protein-coupled receptors. However, histamine-induced I_M modulation in HEK293T cells is novel in that subunits in addition to G_q/11 subunits appear to be involved in the modulation of KCNQ2/3 channel currents.