Nitroglycerin-induced chronic migraine model

Previously studies mostly employed nitroglycerin (NTG) with a dose of 10 mg/kg per injection to model acute or chronic migraine [20, 21]. However, this dose is 1000–10,000 times higher than that used clinically, which caused a drastic and prolonged of the blood pressure drop [22]. In contrast, a previous study has shown that a naturalistic dose of NTG, roughly 8 times higher than the reasonable pharmacological dose used clinically, is sufficient to induce allodynia in rats, without casting doubts that the animal behaviors could be confounded by the abnormal hemodynamic alterations [23, 24]. Thus, in our study, we tested multiple doses of NTG (10 mg/kg, 1 mg/kg and 0.1 kg/mg) to explore the minimal doses required to induce cranial and hindpaw allodynia in mice with least hemodynamic impact. After identifying the ideal dose (0.1 mg/kg) (see results), the mice were intraperitoneally (i.p.) injected with NTG once to mimic acute migraine or every second day for 9 days (i.e., a total of 5 NTG injections) to simulate chronic migraine. The periorbital and hindpaw mechanical thresholds of the mice were tested before and 2 hours after NTG administration on each test day. NTG solution was prepared from a stock solution of 2 mg NTG in 99% propylene glycol (#1466506, Sigma) and freshly diluted in 0.9% saline to a dose of 0.1, 1 and 10 mg/kg. The vehicle controls in these experiments contain 1% propylene glycol diluted in 0.9% saline, used as vehicle compares to that used for three different NTG doses. All injections of NTG were administered as a 0.1, 1 and 10 mg/kg volume via i.p. injection. To avoid confounding, the drugs or vehicles were prepared in a blinded fashion. To evaluate the predictive validity of the model, the migraine specific treatment sumatriptan (#S1198, Sigma), a 5-HT1B/1D agonist, was freshly dissolved in normal saline and i.p. injected at a concentration of 0.6 mg/kg, 5 mins after the NTG injection. The mechanical thresholds were only tested on the 1st, the 9th and the 10th day. These mice were similar to that in mice receiving repeated measurement in other groups [21].