PACAP increases excitability of guinea pig cardiac neurons, an effect mediated by PAC₁ receptors. In dissociated guinea pig cardiac neurons, PACAP causes a positive shift of the voltage dependence of activation of the hyperpolarization-activated, nonselective cation current (I_h). This observation suggested that an enhancement of I_h contributed to the increase in excitability in neurons within cardiac ganglia whole mounts. To evaluate the role of I_h in the PACAP-induced increase in excitability, the increase in action potential generation by 10 nM PACAP was compared in control neurons and neurons treated with 10 or 100 µM ZD7288 or 2 or 2.5 mM cesium chloride (CsCl), drugs known to inhibit I_h. In control cells exposed to PACAP, 1 second depolarizing current pulses elicited multiple action potential firing in 79% of the neurons. In ZD7288 or CsCl containing solutions, the 10 nM PACAP-induced increase in excitability was markedly suppressed, with 7% and 21% of the neurons generating multiple action potentials, respectively. Prior results indicated PACAP initiates depolarization by activating an inward current, which is separate from its enhancement of I_h. Here, we show that a PACAP-induced depolarization was comparable in control neurons and neurons bathed in a CsCl containing solution; an observation indicating that CsCl did not interfere with activation of the PAC₁ receptor by PACAP. Additional experiments indicated that pretreatment with the putative M-current inhibitor, 1 mM BaCl₂, but not 10 µM XE991, initiated multiple firing in a majority of neurons with resting potentials maintained at ~-60 mV. Furthermore, in Ba²⁺-treated cells, 10 nM PACAP increased the number of action potentials generated. Our results indicate that a PACAP enhancement of I_h, rather than an inhibition of I_M and other 1mM Ba²⁺-sensitive potassium currents, is a key ionic mechanism contributing to the peptide-induced increase in excitability for neurons within cardiac ganglia whole mount preparations.