Enhancement of I_h, but not inhibition of I_M, is a key mechanism underlying the PACAP-induced increase in excitability of guinea pig intrinsic cardiac neurons

Abstract

Pituitary adenylate cyclase-activating polypeptide (PACAP) increases excitability of guinea pig cardiac neurons, an effect mediated by PACAP-selective PAC(1) 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 whole-mount cardiac ganglia preparations. To evaluate the role of I(h) in the PACAP-induced increase in excitability, we compared the increase in action potentials generated by 10 nM PACAP in control neurons and in neurons treated with ZD7288 (10 or 100 muM) or CsCl (2 or 2.5 mM), drugs known to inhibit I(h). In control cells exposed to PACAP, 1-s 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 that 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 PAC(1) receptor by PACAP. Additional experiments indicated that pretreatment with the putative M current (I(M)) inhibitor 1 mM BaCl(2), but not 10 microM XE991, initiated multiple firing in a majority of neurons, with resting potentials maintained at approximately -60 mV. Furthermore, in Ba(2+)-treated cells, 10 nM PACAP increased the number of action potentials generated. Our results indicate that PACAP enhancement of I(h), rather than inhibition of I(M) and other 1 mM Ba(2+)-sensitive K(+) currents, is a key ionic mechanism contributing to the peptide-induced increase in excitability for neurons within whole-mount cardiac ganglia preparations.

Nickel suppresses the PACAP-induced increase in guinea pig cardiac neuron excitability

“…Thus, it is imperative to understand what ionic conductances regulate cardiac neuron excitability. Evidence presented previously indicated that a PACAP-enhanced I h was one mechanism contributing to the PACAP-induced increase in cardiac neuron excitability (36,51). A PACAP-induced calcium influx also was found to be a critical step in the peptide modulation of excitability (50).…”

“…2A). This rectification or "sag" in the hyperpolarizing response is due to the activation of I h , the inward current flowing through a hyperpolarization-activated nonselective cationic conductance (HCN) (10,15,36,51,55). Following termination of the hyperpolarization, a hyperpolarization-induced rebound depolarization often occurs (Fig.…”

“…Cardiac neurons are more readily accessible than CNS neurons for experimental manipulations and accordingly, we have used these cells to further our understanding of cellular PAC 1 receptor signaling mechanisms. The results of our previous work indicated that one mechanism contributing to the PACAP/PAC 1 receptor-induced increase in neuronal excitability is via an adenylyl cyclase/cAMP-mediated enhancement of the hyperpolarization-activated, cyclic nucleotide-sensitive nonselective cationic current, I h (36,51). However, results of subsequent studies indicated that application of the potent adenylyl cyclase activator forskolin or exposure to the cellpermeable analog of cAMP 8-bromo-cAMP only partially recapitulated the effect of PACAP on excitability (52), suggesting that PACAP enhancement of other membrane conductances, in addition to I h , also contributes to the peptide-induced modulation of excitability.…”

Pretreatment with Nonselective Cationic Channel Inhibitors Blunts the PACAP-Induced Increase in Guinea Pig Cardiac Neuron Excitability

“…For intracellular recording, atrial whole mount preparations were pinned in a Sylgard-lined chamber and superfused continuously (6–7 ml/min) with Kreb’s solution also containing 10 mM HEPES buffer (Braas et al 1998; Tompkins et al 2006, 2007, 2009; Tompkins and Parsons 2008). The experiments were done with the bathing solution maintained between 32–35°C.…”

Stress peptide PACAP engages multiple signaling pathways within the carotid body to initiate excitatory responses in respiratory and sympathetic chemosensory afferents

“…The PAC1 receptor can also modulate the activity of phospholipase D (PLD) in recombinant cell lines through a small G protein independent of the PLC-PKC pathway (30). Once activated, these pathways do the bidding of PACAP by changing gene expression, releasing Ca 2ϩ from internal stores, and/or activating/phosphorylating a range of ion channels including T-type Ca 2ϩ channels (8,19,38,55,57,58). The cellular mechanisms by which PACAP leads to secretion have been particularly well studied in the rat PC12 cell line, which shares the oxygen sensing and catecholaminergic phenotype of carotid body glomus cells (68).…”