Acetylcholine (ACh) stimulates an increase in potential difference (PD) across intact porcine bronchi due to stimulation of Cl^– and HCO₃^– secretion (Inglis et al 1996). This PD response has been shown to be mediated largely by the M3 muscarinic receptor subtype (Husband & Inglis, 2004) which is expressed along with the M1 subtype in airway submucosal glands (Mak & Barnes, 1990). The aim of this investigation was to study the liquid and mucus secretions produced in porcine bronchi in response to ACh stimulation and investigate the effect of M1 or M3 antagonism on the volume and composition of these secretions. The protocol used was similar to that described previously (Ballard et al. 1999). Cotswold pigs (~15kg) were anaesthetised with inhaled halothane and killed by intravenous overdose of sodium pentobarbitone. The lungs were removed and the bronchi dissected free. Side branches were ligated and the bronchi warmed to 37°C in a tissue bath. The lumen of each bronchus was cleared of liquid and mucus and the tissue was mounted onto a tubular glass support suspended in a beaker of HCO₃^– buffered solution (gassing with 95% O₂-5% CO₂ to maintain pH 7.4) into which the relevant antagonists were added. Antagonists against M1 and M3 subtypes used in these experiments were pirenzipine (PZ) and 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP), respectively. At the end of each experiment all accumulated liquid and mucus was aspirated from within the lumen and transferred to an eppendorf tube. The collected solutions were weighed and dried to distinguish between liquid and solid (mucus) components. Both antagonists significantly reduced the total volume secreted in response to stimulation with 50μM ACh in a concentration dependent manner (Student’s paired t test, P<0.05). 4-DAMP was found to be 106 times more potent than PZ as an inhibitor of liquid secretion with IC₅₀ values of 16.00 ± 1.00 nM (n = 25; means ± S.E.M.) and 17.00 ± 1.86 μM (n = 38), respectively. This suggests the M3 subtype is responsible for mediating liquid secretion from the submucosal glands since PZ is relatively non-specific for the M1 receptor at these concentrations. 4-DAMP pre-treatment also significantly reduced the solid component of secretion (IC₅₀ = 13.00 ± 1.02 nM, n = 25) while the response to PZ could not be plotted sigmoidally. This suggests the M3 subtype is responsible for mediating mucus secretion with M1 playing a minor role if any. In conclusion, ACh-evoked stimulation of liquid and mucus secretion in porcine bronchi is mediated primarily by the M3 receptor.