**3.3 Anti-sympathetic action**

The anti-sympathetic action of cilnidipine has been demonstrated in several experimental studies; increases in the heart rate and plasma catecholamine level induced by cold stress, hypotension or spinal nerve stimulation of the rat were effectively suppressed by cilnidipine but not other Ca2+ channel blockers (Uneyama et al., 1999a). Importantly, the N-type Ca2+ channel-blockade hardly affects parasympathetic neurotransmission (Konda et al., 2001). We compared effects of 4 Ca2+ channel blockers (nifedipine, amlodipine, azelnidipine and cilnidipine) on hypotension-induced sympathetic activation in the halothane-anesthetized canine model (Fig. 2, Takahara et al., 2007; Ishizaka et al., 2010). Intravenous infusion of nifedipine or amlodipine decreased the mean blood pressure with increments of heart rate and cardiac contractility. Azelnidipine also deceased the mean blood pressure with an increment of cardiac contractility whereas marked tachycardia was not induced, which is in accordance with a previous report showing its direct suppressive effects on sinus nodal automaticity. On the other hand, no significant change in the heart rate or cardiac contractility was observed after intravenous infusion of cilnidipine. These results strongly indicate that cilnidipine is desirable among Ca2+ channel blockers to minimize reflex sympathetic nerve activation.

Similar actions of cilnidipine can be observed in the clinical investigations using a parameter of plasma/urinary norepinephrine, 123I-metaiodobenzylguanidine (MIBG), muscle sympathetic nerve activity, low frequency/high frequency ratio (LF/HF ratio) or heart rate, each of which help us better understand their clinical effects on sympathetic nerve activity (Takahara 2009). More importantly, a clinical study of cold pressor test has shown that cilnidipine decreased plasma level of ß-thromboglobulin, a marker of platelet activation, which may prevent arterial thrombosis formation associated with increased sympathetic tone.

#### **3.4 Renal action**

There are observations that L-type Ca2+ channel blockers, such as verapamil, nicardipine, felodipine and nifedipine, induce renal vasodilation and natriuresis in anesthetized dogs and rats. Similarly, cilnidipine increases the renal blood flow and urinary Na+ excretion without affecting creatinine clearance in dogs (Takahara et al., 1997). The kidney is densely innervated by adrenergic nerve fibers. The renal nerve stimulation releases norepinephrine

In animal examinations, cilnidipine has a slow-onset and long-lasting antihypertensive action in spontaneously hypertensive rats, renal hypertensive rats, DOCA-salt hypertensive rats, and 2-kidney 1-clip hypertensive dogs. In addition, cilnidipine significantly decreases the blood pressure of stroke-prone spontaneously hypertensive rats (Watanabe et al. 1995a)

In clinical studies, the antihypertensive effect of cilnidipine has been demonstrated in hypertensive patients, and also in patients with severe hypertension or with complications such as chronic kidney disease, cerebrovascular disease and diabetes. Cilnidipine has been reported to improve some hypertensive conditions closely associated with sympathetic nerve activation such as morning hypertension, nocturnal hypertension, white-coat

The anti-sympathetic action of cilnidipine has been demonstrated in several experimental studies; increases in the heart rate and plasma catecholamine level induced by cold stress, hypotension or spinal nerve stimulation of the rat were effectively suppressed by cilnidipine but not other Ca2+ channel blockers (Uneyama et al., 1999a). Importantly, the N-type Ca2+ channel-blockade hardly affects parasympathetic neurotransmission (Konda et al., 2001). We compared effects of 4 Ca2+ channel blockers (nifedipine, amlodipine, azelnidipine and cilnidipine) on hypotension-induced sympathetic activation in the halothane-anesthetized canine model (Fig. 2, Takahara et al., 2007; Ishizaka et al., 2010). Intravenous infusion of nifedipine or amlodipine decreased the mean blood pressure with increments of heart rate and cardiac contractility. Azelnidipine also deceased the mean blood pressure with an increment of cardiac contractility whereas marked tachycardia was not induced, which is in accordance with a previous report showing its direct suppressive effects on sinus nodal automaticity. On the other hand, no significant change in the heart rate or cardiac contractility was observed after intravenous infusion of cilnidipine. These results strongly indicate that cilnidipine is desirable among Ca2+ channel blockers to minimize reflex

Similar actions of cilnidipine can be observed in the clinical investigations using a parameter of plasma/urinary norepinephrine, 123I-metaiodobenzylguanidine (MIBG), muscle sympathetic nerve activity, low frequency/high frequency ratio (LF/HF ratio) or heart rate, each of which help us better understand their clinical effects on sympathetic nerve activity (Takahara 2009). More importantly, a clinical study of cold pressor test has shown that cilnidipine decreased plasma level of ß-thromboglobulin, a marker of platelet activation, which may prevent arterial

There are observations that L-type Ca2+ channel blockers, such as verapamil, nicardipine, felodipine and nifedipine, induce renal vasodilation and natriuresis in anesthetized dogs and rats. Similarly, cilnidipine increases the renal blood flow and urinary Na+ excretion without affecting creatinine clearance in dogs (Takahara et al., 1997). The kidney is densely innervated by adrenergic nerve fibers. The renal nerve stimulation releases norepinephrine

**3.2 Antihypertensive action** 

**3.3 Anti-sympathetic action** 

sympathetic nerve activation.

**3.4 Renal action** 

and Dahl salt-sensitive rats (Aritomi et al., 2010).

phenomenon, mental stress and cold stress (Yamagishi, 2006).

thrombosis formation associated with increased sympathetic tone.

and induces renal vasoconstriction, anti-natriuresis and renin secretion via activation of adrenoceptors in the vascular vessels, renal tubular cells and granular cells of the juxtaglomerular apparatus, respectively, which cannot be suppressed by typical L-type Ca2+ channel blockers in several experimental studies (Ogasawara et al., 1993). On the other hand, it has been demonstrated that these responses to renal nerve stimulation are suppressed by natriuretic doses of cilnidipine via its N-type Ca2+ channel blocking property.

Fig. 2. Comparison of effects of Ca2+ channel blockers on hypotension-induced sympathetic activation. Nifedipine, amlodipine, azelnidipine or cilnidipine was intravenously administered to halothane-anesthetized dogs, and changes in mean blood pressure, heart rate and cardiac contractility were observed. The intravenous doses of nifedipine, amlodipine, azelnidipine and cilnidipine used were 3, 200, 70 and 3 µg/kg, respectively. Values are expressed as means±SE. Data are quoted and modified from Takahara et al., 2007 and Ishizaka et al., 2010.

Glomerular filtration is essentially regulated by afferent and efferent arterial tone. Since sensitivity of Ca2+ channel blockers to afferent and efferent arteries varies, Ca2+ channel blockers should be appropriately selected for hypertensive patients with chronic kidney disease. Since the sympathetic nerves are distributed to the afferent and efferent arteries, N-

Dual L/N-Type Ca2+ Channel Blocker: Cilnidipine as a New Type of Antihypertensive Drug 37

changes in blood pressure. It is demonstrated that the cerebral blood flow was maintained regardless of whether blood pressure was decreased by cilnidipine. Furthermore, cilnidipine had the activity to shift downward the lower limit of autoregulation for cerebral blood flow according to the results of the estimation of the lower limit of autoregulation for cerebral blood flow by exsanguination (Watanabe et al., 1995b). Interestingly, an antihypertensive and anti-sympathetic dose of cilnidipine reduced the size of cerebral infarction in the rat focal brain ischemia model in contrast to nilvadipine (Takahara et al., 2004), which is in accordance with previous study using a peptidic N-type Ca2+ channel blocker ω-conotoxin MVIIA. Thus, the results may support that N-type Ca2+ channel activation includes

Pancreatic insulin secretion from β-cells and glucagon secretion from α-cells in the islets of Langerhans are Ca2+-dependent processes initiated by Ca2+ influx probably through N-type Ca2+ channels. In a study using N-type Ca2+ channel α1B-subunit-deficient homozygous knockout mice fed normal diet, there was improved glucose tolerance without any change in insulin sensitivity, and also body weight gain reduced in the mice fed a high-fat diet (Takahashi et al., 2005). In another study with fructose-fed rats, insulin sensitivity was significantly lower than in controls, and insulin resistance improved significantly after cilnidipine treatment (Takada et al., 2001). These imply that N-type Ca2+ channels play a

Clinically, it was revealed that cilnidipine significantly reduced 24-hour urinary catecholamines in hypertensive patients with type 2 diabetes, and thereby may improve insulin resistance (Takeda et al., 1999). Also, it is demonstrated that with cilnidipine treatment in patients with obesity, fasting serum immunoreactive insulin (F-IRI) and insulin resistance index as assessed by homeostasis model assessment (HOMA-R) lowered, and serum dehydroepiandrosterone (DHEA) and serum DHEA-sulfate (DHEA-S) increased

Renoprotective, neuroprotective and cardioprotective effects of cilnidipine have been demonstrated in clinical practice or animal examinations. It is noticed that cilnidipine may

Dihydropiridine derivatives including nifedipine have been reported to act as lipophilic chain-breaking antioxidants; however, there are larger differences in their lipophilicity among dihydropyridines. Lipophilicity of cilnidipine is greater than that of amlodipine, which implies that cilnidipine itself can reduce oxidative stress independently in addition to

Excess reactive oxygen species play an essential role in the development of a variety of renal diseases such as glomerulonephritis and tubulointerstitial nephritis. Indeed, in the kidney, cilnidipine significantly inhibited the increase in NADPH oxidase-derived superoxide

have pleiotropic effects besides N-type Ca2+ channel-blocking action.

pathophysiological process of brain ischemia.

significant role in glucose homeostasis.

**4. Pleiotropic effects of cilnidipine** 

its N-type Ca2+ channel blockade action.

(Ueshiba & Miyachi, 2002).

**4.1 Anti-oxidation** 

**3.7 Metabolic syndrome** 

type Ca2+ channel-blocking activity may be partly associated with control of the glomerular pressure. Indeed, cilnidipine has been demonstrated to dilate both afferent and efferent arteries using the hydronephrotic kidney model of the rat (Konno & Kimura, 2008). Furthermore, in renal injury animal models, cilnidipine reduces glomerular capillary pressure, afferent and efferent arteriolar resistances, urinary albumin excretion, and glomerular volume as well as plasma norepinephrine levels.

In clinical studies, cilnidipine significantly decreased urinary albumin excretion without affecting serum creatinine concentration in hypertensive patients, which is comparable to the angiotensin-converting enzyme inhibitor benazepril. Other studies have shown that the renal protective effect of cilnidipine was greater than that of pure L-type Ca2+ channel blockers. Furthermore, the combination of cilnidipine and valsartan was shown to decrease the albumin/creatinine ratio more markedly than valsartan alone. Recently, the multi-center, open-labeled and randomized trial of Cilnidipine versus Amlodipine Randomized Trial for Evaluation in Renal disease (CARTER) has shown that cilnidipine is superior to amlodipine in preventing the progression of proteinuria in patients with hypertension and chronic renal disease when coupled with a renin–angiotensin system inhibitor (Fujita et al., 2007).

## **3.5 Cardiovascular action**

Since the first generation of Ca2+ channel blockers were known to suppress cardiac functions such as contractility, sino-atrial automaticity, and atrioventricular conduction at vasodilator doses, pharmaceutical companies have developed new Ca2+ channel blockers with higher vascular selectivity in addition to slow kinetics as a new generation. The blood-perfused canine heart preparation is an excellent model to quantitatively determine cardio-vascular selectivity of Ca2+ channel blockers, and many Ca2+ channel blockers were analyzed using this model (Taira, 1987). Cilnidipine has about 10 times more potent coronary vasodilator action and higher vascular selectivity than nifedipine in this heart preparation. An in vivo experimental study has confirmed that cilnidipine shows anti-anginal effects in the vasopressin-induced angina model (Saitoh et al., 2003). A recent study indicates that cilnidipine relaxes human arteries through Ca2+ channel antagonism and increases production of nitric oxide by enhancement of endothelial nitric oxide synthase in the human internal thoracic artery (Fan et al., 2011).

The cardioprotective action of cilnidipine against ischemia has been analyzed in a rabbit model of myocardial infarction, in which cilnidipine decreased the myocardial interstitial norepinephrine levels during ischemia and reperfusion periods, leading to reduction of the myocardial infarct size and incidence of ventricular premature beats (Nagai et al., 2005). Clinically, cilnidipine has been reported to improve left ventricular diastolic function in patients with hypertensive heart disease (Kosaka et al., 2009). These cardioprotective actions are probably associated with suppression of cardiac sympathetic overactivity via blockade of N-type Ca2+ channels and/or anti-oxidative (as described below) effects of cilnidipine, which should be further clarified.

#### **3.6 Cerebrovascular action**

The brain is known to have an autoregulatory capacity that allows cerebral blood vessels to maintain constant cerebral blood flow by dilating or contracting in response to abrupt changes in blood pressure. It is demonstrated that the cerebral blood flow was maintained regardless of whether blood pressure was decreased by cilnidipine. Furthermore, cilnidipine had the activity to shift downward the lower limit of autoregulation for cerebral blood flow according to the results of the estimation of the lower limit of autoregulation for cerebral blood flow by exsanguination (Watanabe et al., 1995b). Interestingly, an antihypertensive and anti-sympathetic dose of cilnidipine reduced the size of cerebral infarction in the rat focal brain ischemia model in contrast to nilvadipine (Takahara et al., 2004), which is in accordance with previous study using a peptidic N-type Ca2+ channel blocker ω-conotoxin MVIIA. Thus, the results may support that N-type Ca2+ channel activation includes pathophysiological process of brain ischemia.
