Mouseox system

As in our previous studies [10 (link);23 (link);24 (link)], carotid arterial oxygen saturation, heart rate, respiratory rate, and pulse distension were measured in conscious mice every other day using the MouseOx system with a collar clip sensor (Starr Life Sciences Corp., Allison Park, PA, USA), in accordance with manufacturer’s instructions. Data were collected for a minimum of 10 seconds (150 data points) per sample.

Respiratory rates were determined using the MouseOx system (Starr Life Sciences Corp, Oackmont, PA). In brief, the neck region of the mice were shaved approximately 24hrs prior to respiratory testing. On the day of testing, a sensor that was connected to the MouseOx system (Rev. 6.3) was placed around the neck and a baseline respiratory depression determined. For the baseline, measurements were taken for 5 sec every 5 min until a stable baseline was obtained for at least 25 min. Mice then received the indicated drug or saline subcutaneously and measurements taken for 5 sec every 5 min over at least 50 min.

All experimental procedures were approved by the Institutional Animal Care and Use
Committee. Animals were housed individually and exposed to a 12-h light/dark cycle with food
and water ad libitum. For terminal electrophysiological recordings, all animals were
anesthetized with an intraperitoneal (ip) injection of 50% urethane (1.2g/kg) and
maintained with supplements (5% urethane) administered intravenously (iv) as
needed.^{35 (link),36 (link)} Depth of anaesthesia was assessed by monitoring pinch withdrawal,
corneal reflex, respiration rate and vibrissae movements to maintain an anaesthetic state
III-3.^{35 (link),37 (link)} The jugular vein was catheterized and trachea intubated for the
purposes of iv infusion and pCO₂ monitoring respectively. In addition, oxygen
concentration, blood pressure, heart rate, and respiration were monitored by a Mouse Ox
system (Starr Life Sciences Corp., Oakmont, PA). Body temperature was monitored with a
rectal thermistor and maintained at 37°C with a circulating-water heating pad. The
rat's head was secured with the dorsal surface positioned horizontally in a
stereotaxic device (Kopf Instruments, Tujunga, CA). A small hole was made in the skull and
expanded with bone rongeurs. The exposed dura mater was opened and the extracellular
recording electrode was advanced into the thalamus.

Mice were anesthetized with an intraperitoneal injection of ketamine/xylazine and inoculated intranasally with one-tenth the median lethal dose of mouse-adapted influenza A/PR/8/34 (H1N1) virus. Morbidity as measured by weight loss was monitored daily after infection. Peripheral oxygen saturation (SpO₂) of conscious mice was measured before and after infection using a MouseOx system (Starr Life Sciences Corp., Allison Park, PA). For TNF-α neutralization, mice received 500µg of anti-TNF-α or isotype control antibodies by intraperitoneal injection on the days indicated. For CD8 depletion, mice received 300µg of anti-CD8a or isotype control antibodies by intraperitoneal injection on days 1 and 4 post-infection. For solTNF-α treatment, 2µg of recombinant mouse solTNF-α was intranasally administered at the time of infection.

All methods were approved by the Palmer Institutional Animal Care and Use Committee. Animals were housed individually and exposed to a 12-hour light/dark cycle with food and water ad libitum. For electrophysiological recordings, 9 adult male Wistar rats (320–460 g; Harlan, Indianapolis, IN) were anesthetized with an intraperitoneal injection of 50% urethane (1.2 g/kg) and maintained with supplement doses (5% urethane) administered intravenously as needed.^{48 (link),49 (link)} Anesthetic state III-3 was maintained by monitoring pinch withdrawal, corneal reflex, respiration rate, and vibrissae movements.^{50 (link)} The jugular vein was catheterized for intravenous infusion. The trachea was intubated for PCO₂ monitoring. Oxygen concentration, heart rate, and respiration were monitored by a MouseOx system (Starr Life Sciences Corp, Oakmont, PA). Body temperature was monitored with a rectal thermistor and maintained at 37°C with a circulating-water heating pad. The rat’s head was secured in a stereotaxic device (Kopf Instruments, Tujunga, CA) with its dorsal surface positioned horizontally. A small hole was made in the skull and expanded with bone rongeurs. The exposed dura was opened, and the extracellular recording electrode was advanced into the thalamus.