Powerlab 8 30 system

Inside each of the four experimental chambers electrodes made of stainless steel were used to detect bioelectric potentials in the water generated from the cardioventilatory muscle activity of the fish. A grid placed on the bottom of the chamber acted as one of the electrodes and a rectangle of fine stainless steel wires (2 mm diameter) placed immediately below the water surface constituted the other electrode (figure 2A). A common electric ground electrode was placed in the surrounding water. The fish could move freely between the two electrodes and the electric ground electrode without noticeable effects on the quality of the signal. The raw signals were amplified using four BIO Amplifiers (model ML136, ADInstruments, Castle Hill, Australia) and saved at a frequency of 200 Hz. The BIO Amplifier was pre-set to the following configuration: range: EEG mode, 1 mV; low-pass filter: 120 Hz; high-pass filter: 1 s and with the 50 Hz notch filter activated. The signals from the BIO Amplifiers were directed to a PowerLab 8/30 system (ADInstruments, Castle Hill, Australia) and data were collected on a PC with ADInstruments acquisition software LabChart 7 Pro v7.3.7. In LabChart, the signal was further filtered and processed off-line to separate the electrocardiogram (ECG) signal and ventilatory movements from random muscular activity (see below and figure 2 B–D).

Mice were anesthetized with isoflurane, received low dose buprenorphine for analgesia subcutaneously and were placed on a heating pad to regulate body temperature. Following endotracheal intubation, animals were ventilated with 150 strokes/min and stroke volume of 7 μL/g bodyweight. The left jugular vein was cannulated with PE-10 tubing and a solution of 12.5% bovine serum albumin (Sigma-Aldrich Corp., St. Louis, MO, USA, 2 μL/g bodyweight) was infused. A microtip conductance pressure–volume catheter (1.4F, SPR-839 NR, Millar Instruments, Houston, TX, USA) was inserted into the carotid artery. Heart rate was maintained between 400 and 500 bpm by adjusting the concentration of isoflurane accordingly. The thorax was opened, the heart apex was punctured with a 26 G cannula and another 1.4 F microtip conductance catheter was inserted into the LV. LV pressure and volume and carotid pressure were recorded continuously with an ADInstruments PowerLab 8/30 system (ADInstruments, Spechbach, Germany). Volume calibration was performed using ADInstruments volume calibration cuvette. Cardiac output (CO) was calculated from LV pressure volume loops. Mean arterial pressure (MAP) was calculated from carotid pressure.

In vivo hemodynamic measurements were performed in separate groups at various time points after PVB (3, 6, 9, and 12 weeks). Rats were anesthetized using isoflurane (3% v/v) and placed on a controlled heating table, and the core temperature, measured via rectal probe, was maintained at 37 °C. Left ventricular systolic pressure (LVSP) was measured by catheterizing the left ventricle via the right carotid artery. Right jugular vein access was used for RV catheterization to measure RV systolic pressure (RVSP). Hemodynamic measurements were performed using a 2F Mikro-Tip^® catheter (SPR-320, Millar Instruments, Houston, TX, USA) and a PowerLab 8/30 System with Chart 7.0 software (AD Instruments GmbH, Spechbach, Germany). Immediately after the completion of the hemodynamic measurements, the rats were exsanguinated under deep isoflurane anesthesia (5% v/v, Isofluran Baxter^®, Baxter Deutschland GmbH, Unterschleißheim, Germany), and blood samples were collected.

In vivo hemodynamic measurements were performed after completion of the protocols to induce RV hypertrophy (n = 10 mice per group for each time point). Mice were anesthetized using isoflurane (1.5% v/v) and placed on controlled heating and the core temperature, measured via rectal probe, was maintained at 37°C. The right jugular vein was used for catheterization of the RV to measure RV systolic pressure (RVSP), RV end‐diastolic pressure (RVEDP), and time constant of isovolumic pressure decay Tau. Systemic arterial pressure (SAP) was measured by catheterizing the right carotid artery. Hemodynamic measurements were performed using a Millar microtip catheter (SPR‐671, FMI, Foehr Medial Instruments GmbH, Seeheim/Ober‐Beerbach, Germany) and a PowerLab 8/30 System with the Chart 7.0 Software (AD Instruments GmbH, Spechbach, Germany).

The experiments were conducted as described in [27 (link)]. Briefly, an internodal cell of N. obtusa was placed in a custom-made plexiglass chamber consisting of 14 compartments filled with a solution (APW or a drug dissolved in APW). The compartments were electrically isolated with Vaseline. Two pairs of silver wires were used as extracellular electrodes. Each electrode was placed in a different compartment at a distance of at least 2 cm between adjacent wires. The signal was amplified and digitized using the PowerLab 8/30 system and LabChart 7 software (AD Instruments, Dunedin, New Zealand). Registration began immediately after 10-min incubation. APs were induced electrically (1 µA current stimulus lasting ~0.5 s) in a 10-min period using Stimulus Isolator A365 (WPI, Sarasota, FL, USA) unless the chemical compound induced spontaneous APs. In each case, the third AP of the induced AP series was used for analysis to ensure that the cells were in a stationary state. Conduction velocity was calculated from the time lag between two the same polarity peaks of a conducted AP and the distance between the corresponding recording electrodes. The registration lasted for 3–4 hours unless the cells became unexcitable sooner.