Figure 7.

Endocytosis is important for adaptation of ORNs. A, A 2 h pretreatment of mice with Henkel 100 mixture results in significantly reduced amplitude of EOG responses. Twelve mice were pretreated with Henkel 100 mixture for 2 h, and 11 mice did not receive any treatment (control group). The septal bone with the intact olfactory epithelium was dissected from the head and used intact for EOG recording as described in Materials and Methods. Increasing concentrations of Henkel 100 mixture were applied in the vapor phase for 2 s, with 1 min interval. Bars are the mean amplitude of the EOG responses from the control group (shown in white) and Henkel 100 pretreated mice (shown in black). Factorial ANOVA was performed. Error bars represent SEM; *p < 0.001. B, Treatment of mouse olfactory epithelium with Henkel 100 mixture enhances uptake of fluid phase marker HRP. Olfactory epithelium was dissected from the septal bone and incubated in solution containing 0.25 mg/ml HRP in Ringer’s solution (control; squares) or in Ringer’s solution containing Henkel 100 mixture at 1:10,000 dilution (circles) for indicated periods of time at room temperature. After the incubation, the olfactory epithelium was washed and lysed, and HRP activity was determined as described in Material and Methods. Each point is the mean from three independent experiments. Error bars represent SEM. C, D, Inhibition of clathrin-mediated endocytic pathway affects sensory adaptation in ORNs. C, A representative ratiofluorometric recording of the cytosolic Ca²⁺ level (fluorescence ratio, F₃₄₀/F₃₈₀, viewed as a function of time) shows the effect of dynamin inhibitory peptide (5 min incubation; DIP), resulting in a significant loss of adaptation. Arrows indicate repeated stimulation with a complex odorant mixture (Henkel 50 B, 5 s each) at changing intervals. D, Fluorescence image of an acutely dissociated ORN loaded with the ratiometric Ca²⁺ dye fura-2. Neurons are easily identified based on their characteristic morphology. Changes in Ca²⁺ concentration were measured in the dendritic knob region (depicted by the circle). E, Quantification of the observed effects. Bars represent the average residual response normalized to an initial odorant application as a function of the prestimulation interval. Overall, 17 individual ORNs were included in the analysis. Under control conditions, all of these neurons responded with decreasing signal amplitudes to repeated stimulation. At interstimulus intervals, ≤40 s residual response amplitudes were 57.65 ± 2.14% (control; n = 10) and 78.48 ± 8.85% (treatment; n = 6), whereas at intervals, ≤60 s residual amplitudes were 81.77 ± 4.63% (control; n = 5) and 104.13 ± 5.51% (treatment; n = 5). Response amplitudes before and after pharmacological treatment were statistically analyzed using a homoscedastic Student’s t test. Error bars represent SEM; * p < 0.1; **p < 0.01.