Validation of sympathetic ablation of normal murine salivary glands showing no impact on vascularity or physiological parameters. (A) TH staining of normal murine submandibular salivary glands treated with propranolol (n = 6), 6-OHDA (n = 5), or surgical cervical sympathectomy (n = 7, control n = 15). Scale bar: 50 μm. Quantification of TH⁺ nerves in normal salivary glands as the percentage of TH⁺ nerves out of all Tubb3⁺ nerves in the above conditions. (B and C) TH staining of SMG treated with 6-OHDA (B) (week 1: n = 8 control and 4 6-OHDA; week 3: n = 6 control and 3 6-OHDA) and unilateral cervical sympathectomy (sympathetic cervical ganglionectomy [SCGx]) (C) (week 1: n = 5 per group, week 3: n = 3 per group) at early (week 1) and late (week 3) time points. Scale bar: 200 μm. Quantification of TH⁺ nerve density as TH⁺ area over selected area. (D) Body weight (left panel) (week 0–1: n = 6 control, 7 6-OHDA, and 8 SCGx; week 2–3: 3 per group), excised SMG weight (middle panel) (week 1: n = 3 control, 4 6-OHDA, and 5 SCGx; week 3: n = 3 per group) in mice treated with either 6-OHDA or SCGx at week 0, and cardiac output measured by LV ejection fraction at week 3 (right panel) (n = 3 per group). (E) IF of TH and CD31 of sections from each treatment at week 1 and week 3 (left panel). Quantification of CD31⁺ vessel density showed no significant difference for either treatment as compared with control at each time point (week 1: n = 8 control, 4 6-OHDA, and 5 SCGx; week 3: n = 6 control, 3 6-OHDA, and 3 SCGx). (F) (20 μm sections) TH co-localizes with Tubb3 in sham-treated salivary gland and has diminished expression in the sympathectomized gland. VIP and TRPV1 co-localize with TUBB3 and NFH and maintain expression in both sham and sympathectomized mouse salivary glands. Scale bar: 50 μm. Data are representative of two independent experiments (A–C and E) or one experiment (D). P values were determined by one-way ANOVA, Fisher’s LSD test (A, D, and E), or Student’s t test (B and C). *P < 0.05, **P < 0.01, ****P < 0.0001, ns: not significant.