The distribution and function of P2X and P2Y receptor subtypes were investigated on intact or cultured intramural ganglia of the cat urinary bladder by immunocytochemistry and calcium-imaging techniques, respectively. Neurons were labeled by all seven P2X receptor subtype antibodies and antibodies for P2Y₂, P2Y₄, P2Y₆, and P2Y₁₂ receptor subtypes with a staining intensity of immunoreactivity in the following order: P2X₃=P2Y₂=P2Y₄=P2Y₆=P2Y₁₂>P2X₁=P2X₂=P2X₄>P2X₅=P2X₆=P2X₇. P2Y₁ receptor antibodies labeled glial cells, but not neurons. P2X₃ and P2Y₄ polyclonal antibodies labeled ∼95 and 40% of neurons, respectively. Double staining showed that 100, 48.8, and 97.4% of P2X₃ receptor-positive neurons coexpressed choline acetyl transferase (ChAT), nitric oxide synthase (NOS), and neurofilament 200 (NF200), respectively, whereas 100, 59.2, and 97.6% of P2Y₄ receptor-positive neurons coexpressed ChAT, NOS, and NF200, respectively. Application of ATP, α,β-methylene ATP, and uridine triphosphate elevated intracellular Ca²⁺ concentration in a subpopulation of dissociated cultured cat intramural ganglia neurons, demonstrating the presence of functional P2Y₄ and P2X₃ receptors. This study indicates that P2X and P2Y receptor subtypes are expressed by cholinergic parasympathetic neurons innervating the urinary bladder. The neurons were also stained for NF200, usually regarded as a marker for large sensory neurons. These novel histochemical properties of cholinergic neurons in the cat bladder suggest that the parasympathetic pathways to the cat bladder may be modulated by complex purinergic synaptic mechanisms.