Intravenous Administration of Arecoline Induces Biphasic Modulations in Blood Pressure in Anaesthetized Rats

Meiping Deng1, Xiaowen Ye1, Jiashan Wu1, Lijuan Chen1, Xiaoxia Jiang1, Changzheng Zhang1, Peiling Zhou1* and Yi Luo2*

1School of Educational Sciences and Guangdong Provincial Key Laboratory of Development and Education for Special Needs Children, Lingnan Normal University, 29 Cunjing Road, Zhanjiang 524048, Guangdong Province, China.

2Department of Oncology, Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, Hongshan Road Shizi Street 100, Nanjing, Jiangsu 210028, P.R. China.

ABSTRACT

Arecoline is the main pharmacological alkaloid of areca (betel) nut, which is a traditional medical herb widely used in tropical and subtropical countries, and has several cholinomimetic effects with parasympathetic features. This study was aimed to determine the role of arecoline on systemic blood pressure (BP) modulation and the modulatory characteristics. After rats were anaesthetized, saline or arecoline was intravenously administrated, and the systemic BP signals were recorded. We calculated the reaction times, the mean arterial pressure (MAP), the maximum changes in MAP, and the area under the curve (AUC; MAP change relative to the reaction time) due to arecoline stimulations. The results showed that arecoline induced biphasic modulations in BP, including an initial downregulation (Period 1) and a subsequent upregulation (Period 2), with a concentration-dependent prolonging of reaction times, decreased MAP in Period 1 and increased MAP in Period 2, and elevated maximum changes in MAPs and AUCs. This study provides important evidence that arecoline causes biphasic modulations on systemic BP, which provide basic data for future investigations on the pharmacological characteristics and mechanisms of arecoline action, and raise great concerns regarding the cardiovascular effects of arecoline treatment in clinical practice.

Article Information

Received 01 September 2020

Revised 15 October 2020

Accepted 05 November 2020

Available online 06 January 2021

(early access)

Published 12 November 2021

Authors’ Contribution

MD, XY, JW, LC and XJ performed the experiments and analyzed the data. CZ wrote the manuscript and helped in methodology. PZ and YL designed and conceived the study.

Key words

Arecoline, Blood pressure, Biphasic modulation, Intravenous administration, Anaesthetization, Mean arterial pressure

DOI: https://dx.doi.org/10.17582/journal.pjz/20200901140902

* Corresponding author: peilingzhouvip@163.com, robertluoyi@126.com

0030-9923/2022/0001-0057 $ 9.00/0

Copyright 2022 Zoological Society of Pakistan

Introduction

Arecoline (molecular formula shown in Fig. 1) is a major pharmacological alkaloid of areca (betel) nut, a chewable fruit endemic to South and Southeast Asia that is reported to produce effects of anti-depression, anti-fatigue, attention-focusing, and relaxation (Volgin et al., 2019). Areca nut also acts as a traditional herbal medicine widely used for vermifuge and as a digestant in tropical and subtropical countries (Peng et al., 2015). Arecoline has several cholinomimetic effects on the parasympathetic nervous system (Dasgupta et al., 2017, 2018). Because of the cholinergic features, arecoline is a therapeutic treatment for patients with Alzheimer’s dementia (AD), and ameliorates the symptoms of psychosis and schizophrenia (Bales et al., 2009; Christie et al., 1981; Dasgupta et al., 2006; Pomara and Sidtis, 2010; Xu et al., 2019).

 

Notably, cholinergic actions exert significant effects on cardiovascular activities, but with complicated phenotypes. For example, administration of acetylcholine into specific brain regions induced the blood pressure (BP) depressor response (Shafei et al., 2013; Zhang et al., 2016; Zhu et al., 2015), activation of the muscarinic or nicotinic acetylcholine receptors in the rostral or caudal ventrolateral medulla led to BP pressor or depressor responses (Aberger et al., 2001; Kumar et al., 2009). Studies regarding arecoline on BP modulations have also been published. For example, intraperitoneal (i.p.) injection of arecoline produced cardiodepression in rats and dogs (Beil et al., 1986; Dahl et al., 1994), and a meta-analysis demonstrated that chewing areca nut increased the risk of cardiovascular disorders (Peng et al., 2015; Zhang et al., 2010).

This study was designed to determine the effects of arecoline on systemic BP modulations, as well as the modulatory characteristics on such effects to provide basic data for physiopharmacological investigations, and to provide perspectives of the cardiovascular concerns associated with arecoline use in clinical practice.

Materials and Methods

Animals

Young adult male Sprague–Dawley rats (2 months, 240 ± 20 g; n = 30) were obtained from Jinan Pengyue Experimental Animal Breeding Co. LTD (Jinan, China) and used in this study. The animals were housed in a temperature-controlled (25 ± 1°C) environment with a 12/12 h light/dark-cycle with ad libitum food and water. All animal experiments were conducted according to the National Institutes of Health Guide for the Care and Use of Laboratory Animals (NIH Publication No. 80-23, revised in 1996), and were approved by the Academic and Ethics Committee of Lingnan Normal University (LNU20191018). All efforts were made to minimize the number of animals used, as well as their suffering.

Surgical procedures

The BP recording methods followed those detailed in our previous reports (Zhou et al., 2015; Zhu et al. 2015). Briefly, rats were anesthetized with urethane (1.4 g/kg body weight i.p. injection; Sinopharm Chemical Reagent Co. Ltd., Shanghai, China) and processed for cervical surgery using tracheal intubation. The carotid artery pressure was recorded using a catheter (BL-2020; Taimeng Sci-Tec Co., Ltd., Chengdu, China) that was connected to a signal collecting and processing apparatus (BL-420F, Taimeng Sci-Tech Co., Ltd.) via a BP transducer (PT-100, Taimeng Sci-Tech Co., Ltd.).

BP measurements

Once the BP was stable, saline (0.9% NaCl) and arecoline (at 0.06, 0.2, and 0.6 mg/kg/0.2ml) were injected separately (~10 s) into the vein. Repeated injections were administered at ≥60 min intervals, according to the persistent period from our preliminary experiments in cardiovascular recordings and prior reports on behavioral tests and metabolic examinations (Dasgupta et al., 2018; Sarbani et al., 2006), to avoid mutual interference between arecoline administrations. In general, each animal received 3–5 different drug injections, and the BPs that failed to return to basal values (deviated by >15% of the basal levels) were excluded.

The drug effects on BP regulation were considered substance-specific if they were reversible and reproducible. The mean arterial pressure (MAP), maximum decreased MAP (MDMAP), maximum increased MAP (MIMAP), the latency for arecoline-induced BP changes, the BP reaction time for Period 1 (the duration from the onset of BP decrease to recovery), BP reaction time for Period 2 (the duration from the onset of BP increase to recovery), and the area under the curve (AUC; the changes in MAP relative to the reaction time, as calculated using Graph Pad Prism 5.0 software; Graph Pad Software Inc., San Diego, CA) in response to the drug stimulations were analyzed. As no alterations in BP were observed upon saline treatment, the MAPs in Period 1 and Period 2 for the saline treatment group were calculated from two 60-s BP sequences beginning at 10 s and 100 s, respectively. The pre-test MAPs were calculated form a 60-s BP sequence prior to injections.

Statistical analysis

All data are presented as the mean ± standard error. A one-way analysis of variance followed by a Fisher’s least significant difference post-hoc test was conducted for statistical analyses. P values <0.05 were considered significant.

Results

Compared with saline treatment (Fig. 2A), intravenous injection of arecoline caused biphasic BP regulations, including an initial downregulation (Period 1) and a subsequent upregulation (Period 2; Fig. 2B–D). Statistical analyses revealed no significant differences in the latency of BP changes between groups following arecoline treatment (0.06 mg/kg: 7.66 ± 0.66; 0.2 mg/kg: 8.02 ± 0.72; 0.6 mg/kg: 9.02 ± 0.71 s; F2,45 = 0.886, P = 0.419; Fig. 3A). However, a dose-dependent increase in reaction time in Period 1 (0.06 mg/kg: 42.97 ± 2.54; 0.2 mg/kg: 55.47 ± 2.59; 0.6 mg/kg: 135.73 ± 11.10 s; F2,45 = 69.627, P < 0.001; Fig. 3B) and Period 2 (0.06 mg/kg: 251.41 ± 17.45; 0.2 mg/kg: 414.74 ± 27.32; 0.6 mg/kg: 773.65 ± 66.81 s; F2,45 = 44.221, P < 0.001; Fig. 3C) was observed in response to arecoline stimulations. The durations of Period 2 were 5–8 fold longer than those of Period 1. The overall BP changes recovered within 15 min (Fig. 3B and C; seen also in Fig. 6A), which was consistent with previous reports of arecoline-induced behavioral changes and serum arecoline examinations (Beil et al., 1986; Dasgupta et al., 2018; Pan et al., 2017).

 

 

The MAPs were not significantly different between groups before treatment (saline: 107.23 ± 1.65, 0.06 mg/kg: 109.71 ± 1.17; 0.2 mg/kg: 109.69 ± 1.69; 0.6 mg/kg: 108.91 ± 1.84 mmHg; F3,58 = 0.507, P = 0.679; Fig. 4), while a significant dose-dependent decrease was observed in Period 1 (saline: 107.89 ± 1.86, 0.06 mg/kg: 82.62 ± 1.20; 0.2 mg/kg: 66.50 ± 1.60; 0.6 mg/kg: 62.72 ± 2.44 mmHg; F3,58 = 127.061, P < 0.001; Fig. 4), and a significant increase was observed in Period 2 (saline: 108.27 ± 1.64, 0.06 mg/kg: 117.01 ± 1.15; 0.2 mg/kg: 122.31 ± 1.87; 0.6 mg/kg: 124.40 ± 1.94 mmHg; F3,58 = 16.597, P < 0.001; Fig. 4). Statistical analyses also revealed that the percentage of MAP changes in Period 1 was more intense (~3-fold) than those observed in Period 2.

 

 

Moreover, MDMAP in Period 1 and MIMAP in Period 2 exhibited significant dose-dependent effects (MDMAP: 0.06 mg/kg 47.20 ± 1.99, 0.2 mg/kg 63.25 ± 3.42, 0.6 mg/kg 80.94 ± 4.34 mmHg, F2,45 = 23.318, P < 0.001, Fig. 5A; MIMAP: 0.06 mg/kg 15.31 ± 0.83, 0.2 mg/kg 29.02 ± 1.50, 0.6 mg/kg 39.47 ± 2.42 mmHg, F2,45 = 52.027, P < 0.001, Fig. 5B), which were more drastic (2-3 fold) in MDMAP than MIMAP. Furthermore, the total AUC (the changes in MAP relative to the reaction time) also exhibited significant dose-dependent increases (0.06 mg/kg: 1186.34 ± 65.31, 0.2 mg/kg: 3093.05 ± 250.96, 0.6 mg/kg: 6987.85 ± 327.44; F2,45 = 141.410, P < 0.001; Fig. 6A1–A3 and 6B1), which included dose-dependent increases in the BP downregulation (0.06 mg/kg: 461.09 ± 33.12; 0.2 mg/kg: 973.81 ± 86.62; 0.6 mg/kg: 2460.15 ± 160.73; F2,45 = 104.797, P < 0.001; 6B2) and more intense (~2 fold relative to the downregulation) increases in BP upregulations (0.06 mg/kg 725.76 ± 68.67; 0.2 mg/kg 2120.27 ± 193.57; 0.6 mg/kg 4529.15 ± 340.87; F2,45 = 73.566, P < 0.001; 6B3).

 

Discussion

Arecoline is the main pharmacological alkaloid from areca nut (Dasgupta et al., 2017), and is a therapeutic drug for various ailments, and in particular, for parasitic diseases, digestive disorders, depression, and potentially for AD and schizophrenia treatments (Bales et al., 2009; Chandra et al., 2008; Pomara and Sidtis, 2010). Arecoline primarily exhibits parasympathomimetic features and cardiovascular actions (Beil et al., 1986; Chiou and Kuo, 2008).

The present study showed that intravenous injection of arecoline induced systemic BP modulations with dose-dependent effects. Such effects were reversible and reproducible (Fig. 2), indicating that arecoline exerts substance-specific roles on the regulation of BP. Our current findings demonstrate that arecoline evokes biphasic BP regulations in rats, with an initial downregulation and a subsequent upregulation. However, some reports demonstrated that arecoline only reduced BP in dogs and humans (Beil et al., 1986; Dahl et al., 1994), or a sole BP elevation in rats (Barnes and Roberts, 1991). The discrepancies between the current and previous studies may be due to differences in arecoline doses (a low dose of arecoline may exhibit a primarily depressor response) or sampling times (arecoline induces a transitory BP depression following a longer BP elevation phase; Figs. 3 and 6). Remarkably, the latency of arecoline-induced BP changes via intravenous injection is within 10 seconds in the current study (Fig. 3A), while the onset of cardiovascular activities following areca nut chewing was ~2 minutes (Chu, 2002); thus indicating that different routes of arecoline administration may modulate blood pressure modulation differently.

As arecoline can cross the blood brain barrier (Soncrant et al., 1989), arecoline-mediated BP modulations could be exerted through the central (the brain) and peripheral (mainly the cardiovascular) nervous systems. The downregulation in PB may be due to arecoline activation of the vagal neural circuit, which relaxes the aorta endothelium and improves vasorelaxation (Liu et al., 2016); all such factors correlate with BP depressor responses. Conversely, the upregulation may be due to sympathoadrenal responses (Chu, 2002) since arecoline can activate the hypothalamic-pituitary-adrenal (HPA) axis, which stimulates adrenocorticotropic hormones and corticosterone release (Calogero et al., 1989); thus, leading to a pressor response as proven in many reports (Núñez et al., 2008; Scoggins et al., 1983). Such hypotheses are also supported by evidence that chewing areca nut promotes cadiovascular activities that are associated with sympathoadrenal activation (Chu, 1993, 2002). However, determinations of the exact mechanisms require further investigations.

In summary, this study found that arecoline induced biphasic BP modulations, including an initial downregulation followed by upregulation. Nevertheless, further investigations are required to determine the exact mechanisms controlling such fluctuations in BP.

Acknowledgements

This work was supported by grants from the National Natural Science Foundation of China (81871062, 81773947), the Natural Science Foundation of Guangdong Province (2019A1515010331), Natural Science Foundation of Guangdong Provincial Education Bureau (NO. 2017KTSCX117), and the Special Foundation for Talents of Lingnan Normal University (ZL1819).

Statement of conflict of interest

The authors have declared no conflict of interest.

Reference

Aberger, K., Chitravanshi, V.C. and Sapru, H.N., 2001. Cardiovascular responses to microinjections of nicotine into the caudal ventrolateral medulla of the rat. Brain Res., 892: 138-146. https://doi.org/10.1016/S0006-8993(00)03250-9

Bales, A., Peterson, M.J., Ojha, S., Upadhaya, K., Adhikari, B. and Barrett, B., 2009. Associations between betel nut (Areca catechu) and symptoms of schizophrenia among patients in Nepal: A longitudinal study. Psychiat. Res., 169: 203-211. https://doi.org/10.1016/j.psychres.2008.06.006

Barnes, J.C. and Roberts, F.F., 1991. Central effects of muscarinic agonists and antagonists on hippocampal theta rhythm and blood pressure in the anaesthetised rat. Eur. J. Pharmacol., 195: 233-240. https://doi.org/10.1016/0014-2999(91)90540-7

Beil, M.E., Goodman, F.R., Shlevin, H.H. and Smith, E.F., 1986. Evaluation of the cardiovascular effects of arecoline in the anesthetized dog. Drug Dev. Res., 9: 203-212. https://doi.org/10.1002/ddr.430090304

Calogero, A.E., Kamilaris, T.C., Gomez, M.T., Johnson, E.O., Tartaglia, M.E., Gold, P.W. and Chrousos, G.P., 1989. The muscarinic cholinergic agonist arecoline stimulates the rat hypothalamic pituitary adrenal axis through a centrally-mediated corticotropin-releasing hormone dependent mechanism. Endocrinology, 125: 2445-2453. https://doi.org/10.1210/endo-125-5-2445

Chandra, J.N.N.S., Manish, M., Sadashiva, C.T., Subhash, M.N. and Rangappa, K.S., 2008. Effect of novel arecoline thiazolidinones as muscarinic receptor 1 agonist in Alzheimer’s dementia models. Neurochem. Int., 52: 376-383. https://doi.org/10.1016/j.neuint.2007.07.006

Chiou, S.S. and Kuo, C.D., 2008. Effect of chewing a single betel-quid on autonomic nervous modulation in healthy young adults. J. Psychopharmacol., 22: 910-917. https://doi.org/10.1177/0269881107083840

Christie, J.E., Shering, A., Ferguson, J. and Glen, A.I., 1981. Physostigmine and arecoline: effects of intravenous infusions in Alzheimer presenile dementia. Br. J. Psychiat., 138: 46-50. https://doi.org/10.1192/bjp.138.1.46

Chu, N.S., 1993. Cardiovascular responses to betel chewing. J. Formos. med. Assoc., 92: 835-837.

Chu, N.S., 2002. Neurological aspects of areca and betel chewing. Addict. Biol., 7: 111-114. https://doi.org/10.1080/13556210120091473

Dahl, R.E., Ryan, N.D., Perel, J., Birmaher, B. and Puig-Antich, J., 1994. Cholinergic REM induction test with arecoline in depressed children. Psychiat. Res., 51: 269-282. https://doi.org/10.1016/0165-1781(94)90014-0

Dasgupta, R., Chatterjee, A., Sarkar, S. and Maiti, B.R., 2017. Arecoline aggravates hypothyroidism in metabolic stress in mice. Arch. Physiol. Biochem., 123: 105-111. https://doi.org/10.1080/13813455.2016.1267228

Dasgupta, R., Saha, I., Maity, A., Ray, P.P. and Maiti, B.R., 2018. Arecoline ameliorates hyperthyroid condition in mice under cold stress. Arch. Physiol. Biochem., 124: 436-441. https://doi.org/10.1080/13813455.2017.1420665

Dasgupta, R., Saha, I., Pal, S., Bhattacharyya, A., Sa, G., Nag, T.C., Das, T. and Maiti, B.R., 2006. Immunosuppression, hepatotoxicity and depression of antioxidant status by arecoline in albino mice. Toxicology, 227: 94-104. https://doi.org/10.1016/j.tox.2006.07.016

Kumar, N.N., Ferguson, J., Padley, J.R., Pilowsky, P.M. and Goodchild, A.K., 2009. Differential muscarinic receptor gene expression levels in the ventral medulla of spontaneously hypertensive and Wistar-Kyoto rats: role in sympathetic baroreflex function. J. Hypertens., 27: 1001-1008. https://doi.org/10.1097/HJH.0b013e3283282e5c

Liu, Y.J., Peng, W., Hu, M.B., Xu, M. and Wu, C.J., 2016. The pharmacology, toxicology and potential applications of arecoline: a review. Pharm. Biol., 54: 1-8. https://doi.org/10.3109/13880209.2016.1160251

Núñez, H., Ruiz, S., Soto Moyano, R., Navarrete, M., Valladares, L., White, A., Pérez, H., 2008. Fetal undernutrition induces overexpression of CRH mRNA and CRH protein in hypothalamus and increases CRH and corticosterone in plasma during postnatal life in the rat. Neurosci. Lett., 448: 115-119. https://doi.org/10.1016/j.neulet.2008.10.014

Pan, H., Huang, L., Li, Y., Zhou, X., Lu, Y. and Shi, F., 2017. Liquid chromatography-tandem mass spectrometric assay for determination of unstable arecoline in rat plasma and its application. J. Chromatogr. B., 1070: 112-116. https://doi.org/10.1016/j.jchromb.2017.10.026

Peng, W., Liu, Y.J., Wu, N., Sun, T., He, X.Y., Gao, Y.X. and Wu, C.J., 2015. Areca catechu L. (Arecaceae): A review of its traditional uses, botany, phytochemistry, pharmacology and toxicology. J. Ethnopharmacol., 164: 340-356. https://doi.org/10.1016/j.jep.2015.02.010

Pomara, N. and Sidtis, J.J., 2010. Alzheimer’s disease. N. Engl. J. Med., 362: 1844-1845. https://doi.org/10.1056/NEJMc1002323

Sarbani, G., Idle, J.R., Chi, C. T., Mark, Z., Krausz, K.W. and Gonzale, F.J., 2006. A metabolomic approach to the metabolism of the areca nut alkaloids arecoline and arecaidine in the mouse. Chem. Res. Toxicol., 19: 818-827. https://doi.org/10.1021/tx0600402

Scoggins, B.A., Whitworth, J.A., Coghlan, J.P., Denton, D.A. and Mason, R.T., 1983. Vasodilator prostanoids and ACTH-dependent hypertension. Lancet, 1: 1275. https://doi.org/10.1016/S0140-6736(83)92720-4

Shafei, M.N., Niazmand, S., Enayatfard, L., Hosseini, M. and Daloee, M.H., 2013. Pharmacological study of cholinergic system on cardiovascular regulation in the cuneiform nucleus of rat. Neurosci. Lett., 549: 12-17. https://doi.org/10.1016/j.neulet.2013.05.046

Soncrant, T.T., Holloway, H.W., Greig, N.H. and Rapoport, S.I., 1989. Regional brain metabolic responsivity to the muscarinic cholinergic agonist arecoline is similar in young and aged Fischer-344 rats. Brain Res., 487: 255-266. https://doi.org/10.1016/0006-8993(89)90830-5

Volgin, A.D., Bashirzade, A., Amstislavskaya, T.G., Yakovlev, O.A., Demin, K.A., Ho, Y.J., Wang, D., Shevyrin, V.A., Yan, D., Tang, Z., Wang, J., Wang, M., Alpyshov, E.T., Serikuly, N., Wappler-Guzzetta, E.A., Lakstygal, A.M. and Kalueff, A.V., 2019. DARK classics in chemical neuroscience: Arecoline. ACS Chem. Neurosci., 10: 2176-2185. https://doi.org/10.1021/acschemneuro.8b00711

Xu, Z., Adilijiang, A., Wang, W., You, P., Lin, D., Li, X. and He, J., 2019. Arecoline attenuates memory impairment and demyelination in a cuprizone-induced mouse model of schizophrenia. Neuroreport, 30: 134-138. https://doi.org/10.1097/WNR.0000000000001172

Zhang, C., Luo, W., Zhou, P. and Sun, T., 2016. Microinjection of acetylcholine into cerebellar fastigial nucleus induces blood depressor response in anesthetized rats. Neurosci. Lett., 629: 79-84. https://doi.org/10.1016/j.neulet.2016.06.063

Zhang, L.N., Yang, Y.M., Xu, Z.R., Gui, Q.F. and Hu, Q.Q., 2010. Chewing substances with or without tobacco and risk of cardiovascular disease in Asia: A meta-analysis. J. Zhejiang Univ. Sci. B., 11: 681-689. https://doi.org/10.1631/jzus.B1000132

Zhou, P., Zhu, Q., Liu, M., Li, J., Wang, Y., Zhang, C. and Hua, T., 2015. Muscarinic acetylcholine receptor in cerebellar cortex participates in acetylcholine-mediated blood depressor response in rats. Neurosci. Lett., 593: 129-133. https://doi.org/10.1016/j.neulet.2015.03.036

Zhu, Q., Zhou, P., Wang, S., Zhang, C. and Hua, T., 2015. A preliminary study on cerebellar acetylcholine-mediated blood pressure regulation in young and old rats. Exp. Gerontol., 63: 76-80. https://doi.org/10.1016/j.exger.2015.02.003