Cholinergic and peptidergic modulation of neuronal excitability in rat intracardiac ganglia

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)


Molecular and Cellular Pharmacology

First Committee Member

David J. Adams - Committee Chair


Neonatal rat intracardiac neurons were electrically accessed using the perforated patch configuration of the patch clamp technique. At 23$\sp\circ$C, under current clamp mode, these neurons fire trains of action potentials in response to depolarizing current pulses. Elevating bath temperature to 37$\sp\circ$C reduced the median number of evoked action potentials and decreased input resistance. The adaptation of action potential firing is mediated by the M-current. Voltage deflections elicited by hyperpolarizing current pulses displayed a time-dependent rectification mediated by the H-current.The local anesthetics, procaine and QX-222, selectively blocked nicotinic ACh receptor-channels. The equilibrium dissociation constant (K$\sb{\rm d}$) was 3 $\mu$M for procaine and 31 $\mu$M for QX-222. The K$\sb{\rm d}$'s and voltage sensitivities for local anesthetic binding suggests that the channel M2 region differs from that of frog and rat skeletal muscle and neuronal $\alpha\sb4\beta\sb2$ ACh receptor-channels.Substance P reversibly attenuated nicotinic ACh-evoked current amplitude in a voltage independent manner, with half maximal inhibition occurring at 46 $\mu$M substance P. This effect was not mediated by cAMP, protein kinase C, or protein phosphorylation, suggesting that substance P may be acting directly on the nicotinic ACh receptor-channel.VIP reversibly potentiated whole-cell ACh-evoked membrane currents in neurons electrically accessed using either the perforated patch or the conventional whole-cell configuration, with half maximal potentiation occurring at 260 pM. VIP potentiation is mediated by a membrane receptor and is blocked by the VIP receptor binding inhibitor L-8-K. Binding of VIP to its receptor activates a PTX-sensitive G protein. VIP potentiates ACh-evoked currents by decreasing nAChR-channel closed times and increasing open-channel probability.Rat intracardiac neurons were found to functionally express M$\sb1$, M$\sb2$, M$\sb3$, and M$\sb4$ muscarinic receptor subtypes. Activation of M$\sb1$ muscarinic receptors decreased K$\sp+$ conductance (M-current), whereas M$\sb2$ and M$\sb3$ receptors increased K$\sp+$ and Cl$\sp-$ conductance, respectively. Stimulation of the M$\sb4$ receptor depressed Ca$\sp{2+}$ channel currents through voltage dependent and independent changes in the activation of these channels. The M$\sb4$ receptor effects are mediated by a PTX-sensitive G protein which inhibits $\omega$-CGTX-sensitive, dihydropyridine-sensitive, and $\omega$-CGTX- and dihydropyridine-insensitive Ca$\sp{2+}$ channel subtypes.


Biology, Neuroscience; Health Sciences, Pharmacology

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