ABC | Volume 114, Nº2, February 2020

Original Article Fleury et al. Epinephrine and felypressin effects Arq Bras Cardiol. 2020; 114(2):295-303 are increased in the blood levels of smoker patients. 8 The vasoactive drugs present, therefore, alter the vascular response of smokers, since the nitric oxide (NO) pathway, adrenergic and cholinergic systems are affected. Although epinephrine is considered the safest vasoconstrictor drug in patients with cardiovascular disease by The American Dental Association, 9 patients receiving betablocker treatment who smoke cigarettes must be treated differently. Felypressin may be a safe alternative vasoconstrictor in such populationce since it is a non-adrenergic vasoconstrictor. There is no reported interaction between cigaretes smoking and vasopressin. A previous study showed reduced hypertensive effect of felypressin in rats treated with atenolol. 10 In order to increase the knowledge about the safety of epinephrine and felypressin, this study attempted to test them isolated in intravascular administration to simulate the maximal error in local anesthesia, providing information for the formulation of safer local anesthetic solutions. Therefore, the aim of this study was to evaluate and compare the effects of direct intravenous injection of epinephrine and felypressin on blood pressure of smoker or non-smoker normotensive, hypertensive and atenolol-treated hypertensive rats. Methods For the present study, all norms for animal research were reviewed and approved by the institutional review board before the experiments (protocol #010/2010). Male Wistar rats weighing 140 to 320 g provided by Faculdade de Odontologia de Bauru facilities were used in all groups of this study. The experimental hypertension method, indirect and direct BP measurements, were performed as previously described 10 . All rats received a normal diet, free water and food access and were submitted to 12 hour light/dark cycle. Rats weighing 110 to 150 g were anesthetized with an injection of ketamine (50 mg/kg weight, im, Dopalen ® - Sespo Industry and Trade Ltda., Animal Health Vetbrands Division – Jacareí, São Paulo, Brazil) plus xylazine (10 mg/ kg weight, im, Anasedan ® - Sespo). The left renal artery was isolated, and a 0.25 mm gap silver clip was installed around it, and the right kidney was completely removed. 40,000 IU of small-animal antibiotic (Fontoura Wyeth S.A. – São Bernardo do Campo, São Paulo, Brazil) was injected. It is worth mentioning that surgery for clips implantation in the renal arteries was performed in 10 animals per group, that is, 120 rats. However, each groups consisted of 6 animals, since part of the rats did not have systolic blood pressure above 150 mmHg. Therefore, 72 was the total number of animals used in the study and there were no criteria for the definition of this sample, being defined by convenience. Passive Smoking Method Passive smoking was performed based on previous emphysema induction studies 11 . One day after hypertension induction, the rats were intoxicated one time a day (10 cigarettes per exposure period), 6 days a week. The exposure protocol consisted of confining 10 rats in the inhalation chamber during 10min of compressed air ventilation (10 L/min). Indirect and Direct Blood Pressure Measurements 1K1C rats were heated in individual cages containing an electrical resistance and tail pneumatic cuffs were installed and connected to a digital system for indirect blood pressure record (Physiological Pressure Transducer, ADInstruments Pty. Ltd. – Dunedin, Otago, New Zealand). Rats that presented systolic blood pressure equal to or higher than 150 mmHg in the indirect measurement 15 days after clip surgery were accepted in the hypertensive group or treated with atenolol (90 mg/kg/day; Cristália Pharmaceutical and Chemical Products – Itapira, São Paulo, Brazil) administered by gavage in 1mL for 2 weeks. All groups had blood pressure measured directly for 28-35 days after clip surgery, or the equivalent time in the control group: after ketamine/xylazine anesthesia, a saline-filled polyethylene catheter PE-50 (Clay Adams – Franklin Lakes, New Jersey, USA) with an occluded external extremity was implanted in the left carotid artery and in the right jugular vein. The arterial catheter was connected to a pressure transducer coupled to an invasive blood pressure recording system, using appropriate software (Physiological Pressure Transducer; PowerLab 4/30; Chart Pro – ADInstruments Pty. Ltd). The experiments of intravenous injection of vasoconstrictor drugs were performed with anesthetized rats right after catheter implantation. A scheme of the experiments design are detailed in Figure 1. Dose-Response Curves to Epinephrine and Felypressin Exogenous epinephrine (Adren ® -HipolaborFarmacêuticaLtda – Belo Horizonte, Minas Gerais, Brazil) diluted in saline solution was injected at the doses of 80, 160, 320, 640 and 1,280 ng in bolus through tin venous catheter to obtain dose-response curves. Felypressin alone (Dentsply Pharmaceutical, Catanduva, São Paulo, Brazil) was used at doses of 0.125, 0.25, 0.5, 1, 2 and 3mIU. Intravenous injections in random order were performed after a 3-min interval for each response to stabilize blood pressure. Animals were euthanized with intravenous injection of excessive doses of the anesthetic drug thiopental (Thiopentax ® , Cristália – Chemical and Pharmaceutical Products). The following parameters were analyzed using PAM values (PAM = 1/3 Systolic Pressure + 2/3 Diastolic Pressure): minimal hypotensive response, maximal hypertensive response and response duration. Duration was determined using global pressure alterations, since epinephrine has a complex blood pressure response where there is a hypertensive peak followed by hypotensive response and normalization; previous studies have clarified this pattern 10 . Heart rate was recorded 30 s after the injection during one min to avoid bias caused by longer duration or great blood pressure changes since the program used pulsatile pressure to determine these parameter. In order to compare epinephrine and felypressin, doses that corresponded to 1, 2, 4 and 8 local anesthetic cartridges which would have been administered to the rats, were used. The following formula was used: D = 18,000 (for epinephrine*) or 54 mIU (for felypressin*) x 4,286 x 10 -3 (a weight correction from humans (70 kg) to rats (300 g). * total content in a local anesthetic cartridge 296