The aim of this clinical trial was to evaluate the impacts of administration via the intrarectal route (IR) in cats on their heart and respiratory rates, blood pressure, body temperature, and sedation quality compared to the intramuscular route (IM). The intramuscular group (IMG) received 0.003 mg kg−1 dexmedetomidine, 2 mg kg−1 ketamine, and 0.2 mg kg−1 midazolam while the intrarectal group (IRG) protocol was 0.003 mg kg−1 dexmedetomidine, 4 mg kg−1 ketamine, and 0.4 mg kg−1 midazolam. Cardiorespiratory values, temperature, and sedation score were measured 2 min after administration and then every 5 min up to the 40th minute. Cats belonging to IRG reacted less strongly to the drug, as opposed to those receiving intramuscular administration (2/10 in IRG vs. 8/10 in IMG). Average time between drug administration and standing position was 44.9 ± 5.79 in IRG and 57 ± 9.88 min in IMG. In IRG, maintenance of SpO₂ values is > 95% at each time point. Median and range peak of sedation {7 (5)} in IMG occurs at 20th, 25th, and 30th minutes post drug administration while was lower in IRG. Cardiorespiratory values were slightly lower in IMG than in IRG, but always constant in both treatments. Temperature did not differ between groups. At this dosage, although sedation score was higher in IMG, intrarectal route could be efficacious for performing minimally invasive clinical and diagnostic procedures in cats.