Prostaglandins are important mediators of pain and inflammation. We have examined the effects of prostanoids on voltage-activated calcium currents (I_Ca) in acutely isolated mouse trigeminal sensory neurons, using standard whole cell voltage clamp techniques. Trigeminal neurons were divided into two populations based on the presence (Type 2) or absence (Type 1) of low voltage-activated T-type I_Ca. The absence of T-type I_Ca is highly correlated with sensitivity to µ-opioid agonists and the VR1 agonist capsaicin. In both populations of cells, high voltage-activated I_Ca was inhibited by PGE₂ with an EC₅₀ of about 35 nm, to a maximum of 30 %. T-type I_Ca was not inhibited by PGE₂. Pertussis toxin pre-treatment abolished the effects of PGE₂ in Type 2 cells, but not in Type 1 cells, whereas treatment with cholera toxin prevented the effects of PGE₂ in Type 1 cells, but not in Type 2 cells. Inhibition of I_Ca by PGE₂ was associated with slowing of current activation and could be relieved with a large positive pre-pulse, consistent with inhibition of I_Ca by G protein ß subunits. Reverse transcription-polymerase chain reaction of mRNA from trigeminal ganglia indicated that all four EP prostanoid receptors were present. However, in both Type 1 and Type 2 cells the effects of PGE₂ were only mimicked by the selective EP₃ receptor agonist ONO-AE-248, and not by selective agonists for EP₁ (ONO-DI-004), EP₂ (ONO-AE1-259) and EP₄ (ONO-AE1-329) receptors. These data indicate that two populations of neurons in trigeminal ganglia differing in their calcium channel expression, sensitivity to µ-opioids and capsaicin also have divergent mechanisms of PGE₂-mediated inhibition of calcium channels, with Gi/Go type G proteins involved in one population, and Gs type G proteins in the other.