Asthma is the most common chronic respiratory disease treated with inhaled therapy¹. However, aerosol deposition patterns are complex and imaging methods are needed to improve delivery efficiency. 3D UTE-MRI combined with aerosolized Gd-DOTA had been formerly applied onto spontaneous nose-only-breathing animals². Here, a mechanical administration system was developed to ventilate and nebulize rats. Resulting aerosol distribution and kinetics were compared with free-breathing in healthy and asthmatic animals.

Animal model: Four female Wistar rats (262±2g) were sensitized to ovalbumin (OVA; 1mg/rat)³. Sensitization was verified by plethysmography following OVA-challenge with an increase of the enhanced pause indicator (PenH) correlated to airway resistance⁴. Before imaging, asthmatic rats were challenged by 20-minute OVA-nebulization. Asthmatic symptoms appeared 2h-post and lasted >2h. Three healthy rats (277±4g) were used as controls.

Administration protocol was carried out with a Small Animal Gas Administration System (SAGAS) (Fig.1), compatible with MRI, micro-SPECT, and micro-CT. Two glass syringes (2–6mL/s), actuated by piezomotors, yielded accurate volume control. Chronograms of mouth pressure and expiration mass flow rate were recorded with a Matlab® interface. Breathing cycles and ventilation volumes were set through the same interface. Mechanical ventilation was tuned to match individual free-breathing patterns (45-60 breath/min). Nebulization was synchronized to the animal inspiration.

Imaging protocol was implemented on a clinical 1.5T [Philips Achieva] with a 47mm-diameter receiver coil². Rats, in supine position, were tracheotomized and anaesthetized (isoflurane-O₂) through SAGAS with an intratracheal catheter. Two 3D radial acquisitions (TE₁/TE₂=0.4/1.4ms) were used for R₂^* mapping (pre- and 1h post-nebulization); 3D isotropic T₁W-UTE radial acquisitions (TR/TE=14/0.4ms, α=30º, (0.5mm)³ voxel, T_acq=7.5min) were performed once before, then continuously repeated during nebulization and up to ~1h post-nebulization. Gd-DOTA solution [Dotarem; Guerbet] was nebulized on line [Aeroneb Solo; Aerogen] during ~35min to the rats. A respiration-synchronized cine sequence was used to estimate tidal volume to verify ventilated volume.

Image analysis: Lungs were segmented by a histogram-based method². Relative signal enhancement (RSE) was extracted at each time point and RSE at 15min and at the end of nebulization was converted into concentration maps⁵, with r₁=3.7mM^-1∙s^-1 and baseline T_1,0=1.1s for rat lung at 1.5T. Aerosol distribution was evaluated in the lungs using relative dispersion RD (ratio of standard deviation to mean). Regional distribution at 15min nebulization was assessed by relative concentration deposited in 2 equal volumes (V₁=V₂) divided along 4 anatomical directions (Fig.3). Group analysis was expressed as mean ± standard error of the mean (SEM). Unpaired two-tail t-tests with a significance level of 0.05 were performed.

Baseline signal-to-noise-ratio (SNR) under mechanical ventilation was reduced ~27% in the lung and ~11% in the surrounding tissues. After same nebulization duration (15min), compared to free-breathing, mechanically-ventilated healthy rats had comparable pre-R₂^* (525.63±1.40)s^-1, higher RSE (67.86±28.23)% vs. ~50%², higher deposited aerosol concentration (0.26±0.10)mM vs. 0.15mM². RSE dynamics showed important inter-subject variability but similar trend: continuous increase during and even after nebulization whereas clearance could be monitored during free-breathing. Higher RD was observed in 2 healthy rats, unlike more homogeneous distribution under free-breathing. The effective administered Gd dose over the same duration showed a five-fold increase with SAGAS versus free-breathing.

Baseline signal intensity (SI) was comparable and pre-R₂^* was higher (810.02±1.09)s^-1 in mechanically-ventilated asthmatic rats versus controls. After 15min post-nebulization, asthmatic rats had lower RSE (47.36±15.05)% and lower deposited aerosol (0.19±0.05)mM. After ~30min post-nebulization, SI or concentration differences between two groups were not significant. R₂^* at 1h-post did not present significant differences. Asthmatic rats showed highly heterogeneous enhancement, while controls had relatively lower heterogeneity (Fig.2). A positive correlation was observed between concentration RD and PenH in the asthmatic group at 15min post-nebulization (R²=0.66, p<0.05) and at 35min post-nebulization (R²=0.63, p<0.05), which corresponds to the free-breathing case⁶. Healthy rats showed higher heterogeneity not only H/F (same as free-breathing) but also Ce/Pe direction, while asthmatic rats showed mainly Ce/Pe heterogeneity (Fig.3).

Contrast-enhanced 3D UTE was applied to mechanically-ventilated healthy and asthmatic rats. The administration efficiency was improved by inspiration-synchronized intratracheal nebulization. The correlation between aerosol heterogeneity in asthmatic rats and airway resistance PenH was further confirmed in both breathing modalities, suggesting a pathophysiological marker for asthma: heterogeneous aerosol distribution. The rather low-dose and highly-heterogeneous aerosol deposition observed in asthmatic lungs could be related to the induced bronchoconstrictions⁷. SNR reduction was caused by electronic noise and more controlled breathing. Continuous RSE after nebulization may be due to accidental instillations from accumulated aerosol droplets in the catheter during administration. The method here at 1.5T on animals provides an efficient tool to map aerosol deposition in the lungs. It relies on minimal Gd-DOTA doses, which suggests potential clinical applications.