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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