title: Ethanol alters calcium signaling in axonal growth cones.
abstract: Calcium (Ca2+) channels are sensitive to ethanol and Ca2+ signaling is a critical regulator of axonal growth and guidance. Effects of acute and chronic exposure to ethanol (22, 43, or 87 mM) on voltage-gated Ca2+ channels (VGCCs) in whole cells, and KCl-induced Ca2+ transients in axonal growth cones, were examined using dissociated hippocampal cultures. Whole-cell patch-clamp analysis in neurons with newly-formed axons (Stage 3) revealed that rapidly inactivating, low-voltage activated (LVA) and non-inactivating, high-voltage activated (HVA) currents were both inhibited in a dose-dependent manner by acute ethanol, with relatively greater inhibition of HVA currents. When assessed by Fluo-4-AM imaging, baseline fluorescence and Ca2+ response to ethanol in Stage 3 neurons was similar compared to neurons without axons, but peak Ca2+ transient amplitudes in response to bath-applied KCl were greater in Stage 3 neurons and were decreased by acute ethanol. The amplitude of Ca2+ transients elicited specifically in axonal growth cones by focal application of KCl was also inhibited by acute exposure to moderate-to-high concentrations of ethanol (43 or 87 mM), whereas a lower concentration (22 mM) had no effect. When 43 or 87 mM ethanol was present continuously in the medium, KCl-evoked Ca2+ transient amplitudes were also reduced in growth cones. In contrast, Ca2+ transients were increased by continuous exposure to 22 mM ethanol. Visualization using a fluorescent dihydropyridine analog revealed that neurons continuously exposed to ethanol expressed increased amounts of L-type Ca2+ channels, with greater increases in axonal growth cones than cell bodies. Thus, acute ethanol reduces Ca2+ current and KCl-induced Ca2+ responses in whole cells and axonal growth cones, respectively, and chronic exposure is also generally inhibitory despite apparent up-regulation of L-type channel expression. These results are consistent with a role for altered growth cone Ca2+ signaling in abnormal neuromorphogenesis associated with fetal alcohol spectrum disorders.
authors.affiliation: Center for Neuropharmacology and Neuroscience, Albany Medical College (MC-136), 47 New Scotland Avenue, Albany, NY 12208, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Mah
authors.forename: M W
authors.initials: MW
authors.lastname: Fleck
authors.forename: T A
authors.initials: TA
authors.lastname: Lindsley
mesh_terms: D000818:Animals; D015220:Calcium Channels / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects*; D002478:Cells, Cultured; D000431:Ethanol / Q000494:pharmacology*; D020439:Growth Cones / Q000187:drug effects* / Q000502:physiology; D006624:Hippocampus / Q000166:cytology; D015640:Ion Channel Gating; D018408:Patch-Clamp Techniques; D011189:Potassium Chloride / Q000494:pharmacology; D017966:Pyramidal Cells / Q000187:drug effects* / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D015220:Calcium Channels; D000431:Ethanol; D011189:Potassium Chloride