Chorda Tympani Nerve Recording.

Gustatory nerve responses to lingual application of tastants were recorded from the chorda tympani (CT) branch of the facial nerve as previously described (5). Under pentobarbital anesthesia (50–60 mg/kg body weight), the trachea of each mouse was cannulated, and the mouse was then fixed in the supine position with a head holder to allow dissection of the CT nerve. The right CT nerve was dissected free from surrounding tissues after removal of the pterygoid muscle and cut at the point of its entry to the bulla. The entire nerve was placed on an Ag/AgCl electrode. An indifferent electrode was placed in nearby tissue. Neural activities were fed into an amplifer (K-1; Iyodenshikagaku) and monitored on an oscilloscope and audiomonitor. Whole nerve responses were integrated with a time constant of 1.0 s and recorded on a computer using a PowerLab system (PowerLab/sp4; AD Instrument). For taste stimulation, the anterior half of the tongue was enclosed in a flow chamber made of silicone rubber. Taste solutions [100 mM NH₄Cl, 500 mM sucrose, 500 mM maltose, 500 mM glucose, 500 mM fructose, 1 mM SC45674, 100 mM NaCl, 10 mM quinine-HCl, 10 mM citric acid, 100 mM monopotassium glutamate (MPG)] were delivered to the tongue by gravity flow and flowed over the tongue for 30 s. The tongue was washed with distilled water for an interval of ∼1 min between successive stimulation. Only responses from stable recordings were used in data analysis. Recordings were taken before application, after application, and after 10-min washing out the α-glucosidase inhibitors miglitol and voglibose. For the after inhibitor application condition, miglitol (500 μM) or voglibose (10 μM) was applied to the tongue using a saturated cotton swab for 60 s before testing taste stimuli, followed by rinsing the tongue for 30 s with distilled water, followed by application of taste stimuli for 30 s. Integrated whole nerve response magnitudes were measured 5, 10, 15, 20, and 25 s after stimulus onset, averaged, and normalized to responses to 100 mM NH₄Cl to account for mouse to mouse variations in absolute response magnitudes. This relative response was used for statistical analysis. Two-way ANOVA and the post hoc t test were used to statistically evaluate the differences in responses to a series of tastants before treatment with inhibitor, after incubation with inhibitor, and after washout of α-glucosidase inhibitors applied to the tongue. Statistical analyses were performed using the statistical software packages IBM SPSS Statistics (IBM).