9-(tetrahydro-2-furyl)-adenine and cesium-chloride

Hyperpolarization-activated currents (Ih) are present in several neurons of the central and peripheral nervous system. However, Ih in neurons of the vomeronasal organ (VNO) is not well characterized. We studied the properties of Ih in sensory neurons from acute slices of mouse VNO. In voltage-clamp studies, Ih was identified by the characteristic kinetics of activation, voltage dependence, and blockage by Cs+ or ZD-7288, two blockers of the Ih. Forskolin, an activator of adenylyl cyclase, shifted the activation curve for Ih to less negative potentials. A comparison of Ih properties in VNO neurons with those of heterologously expressed hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, together with RT-PCR experiments in VNO, indicate that Ih is caused by HCN2 and/or HCN4 subunits. In current-clamp recordings, blocking Ih with ZD-7288 induced a hyperpolarization of 5.1 mV, an increase in input resistance, a decrease in the sensitivity to elicit action potentials in response to small current injections, and did not modify the frequency of action potentials elicited by a large current injection. It has been shown that in VNO neurons some pheromones induce a decrease in cAMP concentration, but the physiological role of cAMP is unknown. After application of blockers of adenylyl cyclase, we measured a hyperpolarization of 5.1 mV in 11 of 14 neurons, suggesting that basal levels of cAMP could modulate the resting potential. In conclusion, these results show that mouse VNO neurons express HCN2 and/or HCN4 subunits and that Ih contributes to setting the resting membrane potential and to increase excitability at stimulus threshold.

Topics: Adenine; Animals; Biophysical Phenomena; Biophysics; Cesium; Chlorides; Colforsin; Cyclic AMP; Cyclic Nucleotide-Gated Cation Channels; Dose-Response Relationship, Radiation; Drug Interactions; Electric Stimulation; Enzyme Inhibitors; Gene Expression Regulation; Imines; In Vitro Techniques; Membrane Potentials; Mice; Mice, Inbred C57BL; Neurons, Afferent; Patch-Clamp Techniques; Potassium Chloride; Pyrimidines; Vomeronasal Organ

The role of hyperpolarization-activated, cyclic nucleotide-modulated (HCN) channel isoforms and hyperpolarization-activated cation current (Ih) for seizure-related burst firing in thalamocortical (TC) neurons was investigated in a rat genetic model of absence epilepsy [Wistar Albino Glaxo rats, bred in Rijswijk (WAG/Rij)]. Burst discharges in TC neurons locked to seizure activity in vivo were prolonged during blockade of Ih by Cs+ and ZD7288 (4-ethylphenylamino-1,2-dimethyl-6-methylaminopyrimidinium chloride). In vitro analyses revealed a hyperpolarizing shift of half-maximal Ih activation (Vh) in WAG/Rij (Vh = -93.2 mV) compared with nonepileptic controls [August x Copenhagen-Irish (ACI) (Vh = -88.0 mV)]. This effect is explained by a shift of the responsiveness of Ih to cAMP toward higher concentrations in TC neurons from WAG/Rij, as revealed by application of 8-bromo-cAMP and the phosphodiesterase inhibitor IBMX. During blockade of adenylyl cyclase activity, Ih activation was similar in the two strains, whereas the difference in cAMP responsiveness persisted, thereby voting against different ambient cAMP levels between strains. Increasing the intracellular cAMP level and shifting Ih activation led to a change from burst to tonic firing mode in WAG/Rij but not in ACI rats. Furthermore, HCN1 expression was significantly increased on mRNA and protein levels, with no changes in HCN2-4 expression. In conclusion, there is an increase in HCN1 expression in the epileptic thalamus, associated with a decrease in cAMP responsiveness of Ih in TC neurons and resulting impairment to control the shift from burst to tonic firing, which, in turn, will prolong burst activity after recruitment of Ih during absence seizures.

Topics: 1-Methyl-3-isobutylxanthine; 8-Bromo Cyclic Adenosine Monophosphate; Action Potentials; Adenine; Animals; Blotting, Northern; Cerebral Cortex; Cesium; Chlorides; Cyclic Nucleotide-Gated Cation Channels; Disease Models, Animal; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Drug Interactions; Electric Stimulation; Electroencephalography; Enzyme Inhibitors; Epilepsy, Absence; Female; Gene Expression Regulation; Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; Immunohistochemistry; In Situ Hybridization; In Vitro Techniques; Ion Channels; Male; Membrane Potentials; Microinjections; Neurons; Patch-Clamp Techniques; Potassium Channels; Protein Isoforms; Pyrimidines; Rats; Rats, Mutant Strains; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thalamus