title: Regulation of Kv channel expression and neuronal excitability in rat medial nucleus of the trapezoid body maintained in organotypic culture.
abstract: Principal neurons of the medial nucleus of the trapezoid body (MNTB) express a spectrum of voltage-dependent K(+) conductances mediated by Kv1-Kv4 channels, which shape action potential (AP) firing and regulate intrinsic excitability. Postsynaptic factors influencing expression of Kv channels were explored using organotypic cultures of brainstem prepared from P9-P12 rats and maintained in either low (5 mm, low-K) or high (25 mm, high-K) [K(+)](o) medium. Whole cell patch-clamp recordings were made after 7-28 days in vitro. MNTB neurons cultured in high-K medium maintained a single AP firing phenotype, while low-K cultures had smaller K(+) currents, enhanced excitability and fired multiple APs. The calyx of Held inputs degenerated within 3 days in culture, having lost their major afferent input; this preparation of calyx-free MNTB neurons allowed the effects of postsynaptic depolarisation to be studied with minimal synaptic activity. The depolarization caused by the high-K aCSF only transiently increased spontaneous AP firing (<2 min) and did not measurably increase synaptic activity. Chronic depolarization in high-K cultures raised basal levels of [Ca(2+)](i), increased Kv3 currents and shortened AP half-widths. These events relied on raised [Ca(2+)](i), mediated by influx through voltage-gated calcium channels (VGCCs) and release from intracellular stores, causing an increase in cAMP-response element binding protein (CREB) phosphorylation. Block of VGCCs or of CREB function suppressed Kv3 currents, increased AP duration, and reduced Kv3.3 and c-fos expression. Real-time PCR revealed higher Kv3.3 and Kv1.1 mRNA in high-K compared to low-K cultures, although the increased Kv1.1 mRNA was mediated by a CREB-independent mechanism. We conclude that Kv channel expression and hence the intrinsic membrane properties of MNTB neurons are homeostatically regulated by [Ca(2+)](i)-dependent mechanisms and influenced by sustained depolarization of the resting membrane potential.
authors.affiliation: MRC Toxicology Unit, University of Leicester, Leicester LE1 9HN, UK.
authors.forename: Huaxia
authors.initials: H
authors.lastname: Tong
authors.forename: Joern R
authors.initials: JR
authors.lastname: Steinert
authors.forename: Susan W
authors.initials: SW
authors.lastname: Robinson
authors.forename: Tatyana
authors.initials: T
authors.lastname: Chernova
authors.forename: David J
authors.initials: DJ
authors.lastname: Read
authors.forename: Douglas L
authors.initials: DL
authors.lastname: Oliver
authors.forename: Ian D
authors.initials: ID
authors.lastname: Forsythe
mesh_terms: D000200:Action Potentials / Q000502:physiology; D000818:Animals; D001306:Auditory Pathways / Q000502:physiology*; D015153:Blotting, Western; D001933:Brain Stem / Q000502:physiology; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000502:physiology; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D004594:Electrophysiology; D007150:Immunohistochemistry; D015640:Ion Channel Gating / Q000502:physiology; D009474:Neurons / Q000502:physiology*; D009924:Organ Culture Techniques; D010766:Phosphorylation; D011149:Pons / Q000502:physiology*; D011188:Potassium / Q000494:pharmacology; D024642:Potassium Channels, Voltage-Gated / Q000502:physiology*; D016760:Proto-Oncogene Proteins c-fos / Q000096:biosynthesis; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology
chemical_list: D015220:Calcium Channels; D017362:Cyclic AMP Response Element-Binding Protein; D024642:Potassium Channels, Voltage-Gated; D016760:Proto-Oncogene Proteins c-fos; D011188:Potassium; D002118:Calcium