Pulse pulsefit software

Catecholamine secretion from glomus cells in CB slices was performed as described previously in our laboratory (70, 71) . To test responsiveness to hypoxia, hypercapnia, or hypoglycemia, slices were transferred to a recording chamber and continuously perfused with different recording solutions (see recording solutions). Secretory events were recorded with a 10-m carbon fiber electrode. Amperometric currents were recorded with an EPC-8 patch-clamp amplifier (HEKA Elektronik, Lambrecht/Pfaltz, Germany), filtered at 100 Hz, and digitized at 250 Hz before storage on computer. Data acquisition and analysis were performed with an ITC-16 interface (InstruTECH Corporation, NY, USA) and PULSE/PULSEFIT software (HEKA Elektronik). The secretion rate (femtocoulombs per minute) was calculated as the amount of charge transferred to the recording electrode during a given period of time. The cumulative secretion signal was the sum of charges of successive amperometric events during a given time period.

Macroscopic ionic currents were recorded from dispersed mouse glomus cells using the whole-cell configuration as adapted in our laboratory (70, 72) . Patch-clamp pipettes (2 to 3 megohms) were pulled from capillary glass tubes (Kimax, Kimble Products) with a horizontal pipette puller (model P-1000, Sutter Instrument) and fire-polished with a microforge (MF-830, Narishige). Voltage-clamp recordings were obtained with an EPC-7 amplifier (HEKA Elektronik). The signal was filtered (3 to 10 kHz), digitized with an analog/digital converter (ITC-16 InstruTECH Corporation), and sent to the computer. Data acquisition and storage were performed using the PULSE/PULSEFIT software (HEKA Elektronik) at a sampling interval of 20 s.

Adult rat DRG neurons were isolated and cultured according to the method described by Herzog et al. (2003) . Briefly, rats were anesthetized and decapitated. Cells were treated with collagenase (1 mg/mL) and papain (1 mg/mL), dissociated in DMEM supplemented with 10% fetal bovine serum, and plated on glass coverslips coated with poly-L-ornithine and laminin. Cultures were maintained at 37 o C in a 5% CO 2 incubator, and media was changed every 2 days during experimental incubation periods. DRG neurons express both tetrodotoxin-sensitive (TTX-S) and TTX-resistant (TTX-R) sodium channels.
Whole-cell recordings were performed on DRG neurons. Membrane currents were measured with pipettes (2e5 MU) pulled from glass capillary tubes and connected to an EPC-9 amplifier operating Pulse:Pulsefit software (HEKA elektronik, Germany). The pipette solution was composed of (in mM): CsCl 120, TEACl 20, Na 2 ATP 5, EGTA 10, HEPES 10, MgCl 2 2.5, Na 2 GTP 0.4. The pH of the solution was adjusted to 7.3 with CsOH. The external solution was composed of (in mM): NaCl 140, KCl 5, CaCl 2 2.5, MgCl 2 1, HEPES 10, D-glucose 10. The solution was adjusted to pH 7.3e7.4 with NaOH. Ag-AgCl saline bridge was used for the reference electrode. The peak sodium currents were selected out and normalized by their relative amplitude to control.