Title

Voltage-dependent ionic conductances and their modulation by alpha-adrenergic agonists in cultured parasympathetic neurones from rat intracardiac ganglia

Date of Award

1991

Availability

Article

Degree Name

Doctor of Philosophy (Ph.D.)

Department

Molecular and Cellular Pharmacology

First Committee Member

David J. Adams, Committee Chair

Abstract

Voltage-dependent ionic conductances underlying the resting membrane potential (RMP) and action potential of cultured parasympathetic neurons from rat intracardiac ganglia were studied using the whole-cell patch-clamp technique. These neurones have a mean RMP of $-$52 mV and mean input resistance of 850 M$\Omega$. The current-voltage relationship indicated that both leakage and voltage-dependent K$\sp{+}$ currents contribute to the RMP. The rising phase of action potential depended on inward Na$\sp{+}$ and Ca$\sp{2+}$ currents, and the repolarization and after-hyperpolarization were due to outward K$\sp{+}$ current. Depolarization activated outward K$\sp{+}$ current is comprised of a delayed rectifying K$\sp{+}$ current (I$\sb{\rm Kv}$) and a Ca$\sp{2+}$-activated K$\sp{+}$ current (I$\sb{\rm KCa}$). Bath application of charybdotoxin selectively blocked I$\sb{\rm KCa}$, and external TEA$\sp{+}$ blocked both I$\sb{\rm Kv}$ and I$\sb{\rm KCa}$ dose-dependently. Voltage-dependent Na$\sp{+}$ current exhibited kinetics similar to those reported for Na$\sp{+}$ current of rat sympathetic neurones, and was completely and reversibly blocked by tetrodotoxin. Depolarization-activated Ca$\sp{2+}$ currents (I$\sb{\rm Ca}$) exhibited a high threshold for activation with no evidence for the presence of a low-threshold I$\sb{\rm Ca}$ in the soma of rat cardiac neurones. The voltage-dependent kinetics of onset of, and recovery from, inactivation were described by the sum of two exponential functions. At least three distinct types of Ca channels are present in these neurones: dihydropyridine sensitive, $\omega$-conotoxin-sensitive and dihydropyridine- and $\omega$-conotoxin-resistant Ca channels. I$\sb{\rm Ca}$ was modulated by norepinephrine (NE) whereby NE reduced the amplitude and rate of activation of I$\sb{\rm Ca}$. NE inhibition of I$\sb{\rm Ca}$ was mimicked by methoxamine and antagonized by phentolamine, indicating the involvement of $\alpha\sb1$-adrenoceptors. In 35% of neurones studied, NE induced a simultaneous inhibition of I$\sb{\rm Ca}$ and activation of a background current carried by small cations. The inhibition of I$\sb{\rm Ca}$ and activation of background current were mimicked by intracellular GTP-$\gamma$-S, but antagonized either by intracellular GDP-$\beta$-S or by pretreatment of the neurones with pertussis toxin. These results suggest that activation of $\alpha\sb1$-adrenoceptors by NE and the modulation of I$\sb{\rm Ca}$ and background current is coupled by a pertussis toxin-sensitive G-protein(s).

Keywords

Biology, Neuroscience; Health Sciences, Pharmacology; Biology, Animal Physiology

Link to Full Text

http://access.library.miami.edu/login?url=http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:9214846