How are inhalations performed on a rat using a nebulizer? - briefly
The animal is positioned in a sealed exposure chamber linked to a nebulizer that generates a fine aerosol of the test solution at a calibrated flow rate. During a predetermined exposure interval the rat breathes the aerosol spontaneously (or under light restraint), after which the chamber is vented and the animal returned to its housing.
How are inhalations performed on a rat using a nebulizer? - in detail
Delivering aerosolized agents to a laboratory rat with a nebulizer requires a controlled environment, precise equipment setup, and consistent monitoring of respiratory parameters. The following protocol outlines each component of the procedure.
A. Preparation of the animal
- Acclimate the rat to the experimental room for at least 30 minutes to reduce stress‑induced respiratory variability.
- Verify body weight and health status; record baseline respiratory rate and tidal volume using a plethysmograph if available.
- Anesthetize with an inhalational agent (e.g., isoflurane 1–2 % in oxygen) or an injectable regimen (e.g., ketamine/xylazine 80/10 mg kg⁻¹) chosen for minimal impact on airway tone. Maintain anesthesia depth by checking pedal reflex and respiratory pattern.
- Place a temperature‑controlled heating pad beneath the animal to preserve core temperature (≈37 °C).
B. Nebulizer configuration
- Select a jet or ultrasonic nebulizer capable of generating particles with a mass median aerodynamic diameter of 1–5 µm, appropriate for alveolar deposition.
- Connect the nebulizer to a calibrated air‑flow source (typically 1–2 L min⁻¹) equipped with a flowmeter and a bacterial/viral filter downstream of the aerosol chamber.
- Load the solution of interest (e.g., drug, tracer) into the nebulizer reservoir; ensure volume does not exceed the manufacturer’s maximum (often 5 ml).
- Prime the system by running aerosol for 30 seconds to achieve a stable output, then discard the initial aerosol to eliminate dead‑space contamination.
C. Exposure chamber assembly
- Use a sealed, transparent inhalation chamber sized for a single rat (approximately 2 L). The chamber should have inlet and outlet ports allowing continuous airflow through the nebulizer.
- Attach the nebulizer outlet to the chamber inlet; connect the chamber outlet to a suction pump set to maintain a slight negative pressure (0.2–0.5 L min⁻¹) that draws aerosol through the animal’s breathing zone.
- Verify that the chamber interior is free of condensation; if needed, incorporate a desiccant or humidification control to maintain relative humidity between 30–50 %.
D. Execution of the inhalation session
- Transfer the anesthetized rat into the chamber using forceps or a gentle scoop; position the animal so that the snout aligns with the aerosol stream.
- Close the chamber lid and start the nebulizer; record the start time.
- Maintain exposure for the predetermined duration (commonly 5–30 minutes) depending on the dose calculation:
Dose (µg) = Concentration (µg ml⁻¹) × Nebulizer output (ml min⁻¹) × Exposure time (min) × Inhaled fraction (≈0.5). - Monitor respiratory rate, oxygen saturation (if pulse oximetry is available), and anesthesia depth every 5 minutes. Adjust anesthetic delivery or airflow as needed to keep physiological parameters within normal limits.
E. Post‑exposure handling
- After the exposure period, cease nebulization and allow the chamber to clear for 1–2 minutes before opening.
- Transfer the rat to a recovery cage on a warming pad; discontinue anesthetic agents and monitor until the animal regains righting reflex.
- Collect biological samples (blood, lung tissue, bronchoalveolar lavage) at the appropriate time points for downstream analysis.
- Clean the nebulizer and chamber according to biosafety guidelines: rinse with distilled water, disinfect with 70 % ethanol, and allow to dry before the next use.
F. Quality control and documentation
- Record nebulizer performance metrics (particle size distribution, aerosol output rate) for each session.
- Log animal identifiers, anesthesia protocol, exposure duration, and any deviations from the standard procedure.
- Store all data in a laboratory information management system to ensure traceability and reproducibility.
Following this systematic approach yields reproducible aerosol delivery to rodents, enabling accurate assessment of inhaled therapeutics, toxicants, or experimental probes.