Als Breeding pairs of heterozygous Kcna1-null mice on a C3HeB/FeJ congenic background were purchased from Jackson Laboratories (Bar Harbor, Maine) and colonies have been maintained inside the Animal Resource Facilities at Creighton University School of Medicine plus the Barrow Neurological Institute. Mice have been provided meals and water ad libitum and kept on a 12-hour light/dark cycle. All procedures involving animals had been in accordance with National Institutes of Wellness recommendations, the EU Directive 2010/63/EU and have been authorized by the Institutional Animal Care and Use Committees at Creighton University College of Medicine plus the Barrow Neurological Institute St. Joseph’s Healthcare Center. Acute slice preparation Wild-type and Kcna1-null littermates (P30-P45) were anesthetized with isoflurane, decapitated, and their brains removed and promptly placed into ice cold, oxygenated (95 O2/5 CO2) artificial cerebrospinal fluid (aCSF) containing (in mM): 206 Sucrose, 2.eight KCl, 1 CaCl2, 1 MgCl2, 2 MgSO4, 26 NaHCO3, 1.25 NaH2PO4, and 10 glucose (pH 7.four). Horizontal sections (350 ) of ventral hippocampal-entorhinal cortex (HEC) have been cut on a Leica VT1200 (Leica Microsystems Inc., Bannockburn, IL, USA) and transferred to a holding chamber for 1 hr with warm (32 ) oxygenated aCSF containing (in mM): 125 NaCl, 3.0 KCl, two.4 CaCl2, two.5 MgSO4, 26 NaHCO3, 1.25 NaH2PO4, and 10 glucose (pH 7.four) as we’ve got described (Simeone et al., 2011). All chemical substances and drugs had been purchased from Sigma-Aldrich (St. Louis, MO, U.S.A.). Multi-microelectrode recordings Procedures were carried out as we have previously described (Simeone et al., 2011). Briefly, a HEC slice was placed more than the complete electrode grid of a MED64 probe (see Figure 1A; Alpha Med Systems, Osaka, Japan). A custom probe cap allowed delivery of humidified air (95 O2/5 CO2) and perfusion (1 ml min-1) of in-line pre-warmed oxygenated aCSF that resulted inside a solution inlet temperature of 33.2 and an outlet temperature of 31.6 . The bath level was maintained near interface. Spontaneous and evoked extracellular potentials have been recorded with either Conductor v3 (Alpha Med Systems) or Mobius v2 (Witwerx Inc., Tustun, CA, USA) application and acquired at a 20 kHz sampling price having a bandwidth of 0.3-Borono-4-fluorobenzoic acid site 1 Hz?0 kHz.Methyl 6-oxopiperidine-3-carboxylate site Input/output (I/O) curves of presynaptic fiber volleys, postsynaptic field potentials and short-term plasticity were obtained by successive paired stimulations (50 ms inter-stimulus interval) of growing intensity (ten ?180 ) separated by 45 sec.PMID:23290930 The mossy fiber (MF)-CA3, medial perforant path (MPP)-dentate granule cell (DG), MPP-CA3, lateral perforant path (LPP)-DG and LPP-CA3 synapses had been tested in each slice. Signal analysis Information was imported into Spike2 (v6) application (Cambridge Electronic Design, Cambridge, England) as we have previously described (Simeone et al., 2011). Briefly, to recognize DC shifts of the SPWs, raw recordings have been subjected to a 50 Hz low-pass filter (-3dB point = 70 Hz) utilizing a Finite Impulse Response (FIR) filter (1319 filter coefficients) supplied inside the Spike2 computer software and down-sampled to two.five kHz. Automated threshold detection of SPWs was set at 3 times the amplitude on the peak-to-peak noise level. SPW frequencies wereNeurobiol Dis. Author manuscript; readily available in PMC 2014 June 01.Simeone et al.Pagedetermined by fitting inter-SPW interval histograms using a Gaussian function and calculating the weighted Gaussian imply frequency.NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author M.
