GABAergic Mechanism of Propofol Toxicity in Immature Neurons

Abstract

Certain anesthetics exhibit neurotoxicity in the brains of immature but not mature animals. Gamma-aminobutyric acid (GABA), the primary inhibitory neurotransmitter in the adult brain, is excitatory on immature neurons via its action at the GABAA receptor, due to a reversed transmembrane chloride gradient. GABAA receptor activation in immature neurons is sufficient to open L-type voltage gated calcium channels. As propofol is a GABAA agonist, we hypothesized that it and more specific GABAA modulators would increase intracellular free calcium ([Ca2+]i), resulting in the death of neonatal rat hippocampal neurons. Neuronal [Ca2+]i was monitored using Fura2-AM fluorescence imaging. Cell death was assessed by double-staining with propidium iodide and Hoechst 33258 at 1 h (acute) and 48 h (delayed) after 5 h exposure of neurons to propofol or the GABAA receptor agonist, muscimol, in the presence and absence of the GABA receptor antagonist, bicuculline, or the L-type Ca2+ channel blocker, nifedipine. Fluorescent measurements of caspase-3,-7 activities were performed at 1 h after exposure. Both muscimol and propofol induced a rapid increase in [Ca2+]i in day in vitro (DIV) 4, but not in DIV 8 neurons, that was inhibited by nifedipine and bicuculline. Caspase-3,-7 activities and cell death increased significantly in DIV 4 but not DIV 8 hippocampal neuronal cultures 1 h after a 5 h exposure to propofol, but not muscimol, and were inhibited by the presence of bicuculline or nifedipine. We conclude that an increase in [Ca2+]i, due to activation of GABAA receptors and opening of L-type calcium channels, is necessary for propofol-induced death of immature rat hippocampal neurons but that additional mechanisms not elicited by GABAA activation alone also contribute to cell death.

Regional and Temporal Profiles of Calpain and Caspase-3 Activities in Postnatal Rat Brain following Repeated Propofol Administration

“…The conversion of GABA receptors occurs during the first week of postnatal development, and at day 7, some regions have either completed the switch or are well along in the process of switching. Thus, GABAergic excitation is most likely region-dependent and should be considered as a contributor to damage [31] . A further possibility is that the expression of GABA A subunits is different in these regions at this stage of development.…”

“…To resolve whether calpain activity which coincides with caspase-3 activity participates in apoptotic and/or necrotic cell death will need additional data and ultrastructural analyses. Propofol treatment is known to change calcium homeostasis [13,31] that can modulate the activity of calpain, a calcium-sensitive protease. This was confirmed in our study.…”

Propofol Compared with Isoflurane Inhibits Mitochondrial Metabolism in Immature Swine Cerebral Cortex

“…Experimental evidence shows that immature neurons, which exhibit enhanced excitability compared with adults, are particularly vulnerable to caspase activation and hence apoptosis by anesthetics including propofol. 1,5 Disturbances in Ca þ þ homeostasis can lead to neurodegeneration in the developing brain, 6 which shows different mechanisms for depolarization than the adult. 5 Anesthetics used in pediatrics, and in particular propofol, can increase Ca þ þ influx.…”

“…1,5 Disturbances in Ca þ þ homeostasis can lead to neurodegeneration in the developing brain, 6 which shows different mechanisms for depolarization than the adult. 5 Anesthetics used in pediatrics, and in particular propofol, can increase Ca þ þ influx. 7 Thus, enhanced caspase activation by Ca þ þ has been implicated as the mechanism for neuronal death.…”

“…7 Thus, enhanced caspase activation by Ca þ þ has been implicated as the mechanism for neuronal death. 7 Although, experiments performed in vitro show that the rapid rise in Ca þ þ induced by propofol can occur before neuronal apoptosis, 5 this change in flux does not always precede and therefore does not fully explain the anesthetic-induced neurotoxicity.…”