**7. Aminopeptidase A cerebral inhibitors**

Aminopeptidases (AP) are proteolytic enzymes ubiquitously distributed, capable of hydrolyzing the aminoterminal amino acids of peptides and polypeptides that have an important role in controlling them centrally, as well as in peripheral tissues and blood. Their activities reflect the functional state of their endogenous substrates.

The terminology is confusing in relation to aminopeptidases, since the same enzyme is usually identified by different names. Aminopeptidase A (EC 3.4.11.7) hydrolyses the terminal of amino acids, mainly glutamic residues, but is also able to recognize the terminal of aspartic, so this enzyme is also known as aminopeptidase glutamate(GluAP).(Bodineau et al., 2008)

Several aminopeptidases (angiotensinase) are involved in the metabolism of the major active peptides of the RAAS. In particular, they are involved in the metabolism of Ang II, Ang III, Ang IV and Ang 2-10. Ang III is derived from the metabolism of Ang II by the action of the GluAP that hydrolyzes the peptide bond with an acid residue, Asp. The AlaAP and/or the ArgAP metabolizes Ang III to Ang IV by hydrolysis of the amino terminal Arg. The Ang I is transformed to Ang 2-10 by the action of the AspAP after the release of amino terminal Asp. The Ang III is a less potent vasoconstrictor than Ang II. It stimulates adrenal secretion of aldosterone, a neural stimulator and has the same affinity for the AT1 and AT2 receptors. Ang IV has little affinity for the AT1 and AT2 receptors and a lot for the AT4 receptor. Ang IV has an important role in regulating local blood flow, including the brain, but also has been assigned a role in cognitive processes, stress, anxiety and depression. Ang 2-10 opposes the vasoconstrictor effect of Ang II.

However, it is also said that this peptide would lead to aortic contraction dose dependent, through the AT1 receptor. Now, in regard to BP control, working with the hypothesis of a coordinated action of different peptides of the system acting together on the AT1, AT2 and AT4 receptors. Wright and colleagues (Wright & Harding, 1992, Wright & Harding, 1994, Wright & Harding, 1997) analysed the role of brain aminopeptidases in hypertension in several studies. They demonstrated that after intracerebroventricular injection of Ang II and

A new antihypertensive agent, rostafuroxin (PST2238) a digitalis derivative, has been developed due to the ability to correct abnormalities of the Na-K pump. It is endowed with high potency and efficacy in reducing BP and preventing organ hypertrophy in animal models. (Ferrari et al., 2006) At the molecular level in the kidney, rostafuroxin normalizes the increased activity of the Na-K pump induced by adducin mutants pump and endogenous ouabain. In the vasculature, it normalizes the increasement of myogenic tone

A very high safety factor is the lack of interaction with other mechanisms involved in the regulation of BP, along with evidence of high tolerability and efficacy in hypertensive patients point to the rostafuroxin as the first example of a new class of antihypertensive drug designed to antagonize endogenous ouabain and adducin. Phase II clinical trial was recently completed, Ouabain and Adducin for Specific Intervention on Sodium in Hypertension Trial (OASIS-HT), in which rostafuroxin was used with encouraging results as 23% had a significant decrease in BP. In the future we will have to wait to compare the results of rostafuroxin with other antihypertensive drugs to be validated as ACE inhibitors

Aminopeptidases (AP) are proteolytic enzymes ubiquitously distributed, capable of hydrolyzing the aminoterminal amino acids of peptides and polypeptides that have an important role in controlling them centrally, as well as in peripheral tissues and blood. Their

The terminology is confusing in relation to aminopeptidases, since the same enzyme is usually identified by different names. Aminopeptidase A (EC 3.4.11.7) hydrolyses the terminal of amino acids, mainly glutamic residues, but is also able to recognize the terminal of aspartic, so

Several aminopeptidases (angiotensinase) are involved in the metabolism of the major active peptides of the RAAS. In particular, they are involved in the metabolism of Ang II, Ang III, Ang IV and Ang 2-10. Ang III is derived from the metabolism of Ang II by the action of the GluAP that hydrolyzes the peptide bond with an acid residue, Asp. The AlaAP and/or the ArgAP metabolizes Ang III to Ang IV by hydrolysis of the amino terminal Arg. The Ang I is transformed to Ang 2-10 by the action of the AspAP after the release of amino terminal Asp. The Ang III is a less potent vasoconstrictor than Ang II. It stimulates adrenal secretion of aldosterone, a neural stimulator and has the same affinity for the AT1 and AT2 receptors. Ang IV has little affinity for the AT1 and AT2 receptors and a lot for the AT4 receptor. Ang IV has an important role in regulating local blood flow, including the brain, but also has been assigned a role in cognitive processes, stress, anxiety and depression. Ang 2-10

However, it is also said that this peptide would lead to aortic contraction dose dependent, through the AT1 receptor. Now, in regard to BP control, working with the hypothesis of a coordinated action of different peptides of the system acting together on the AT1, AT2 and AT4 receptors. Wright and colleagues (Wright & Harding, 1992, Wright & Harding, 1994, Wright & Harding, 1997) analysed the role of brain aminopeptidases in hypertension in several studies. They demonstrated that after intracerebroventricular injection of Ang II and

this enzyme is also known as aminopeptidase glutamate(GluAP).(Bodineau et al., 2008)

and ARBs and its influence on the control of BP. (Staessen et al., 2011)

activities reflect the functional state of their endogenous substrates.

**7. Aminopeptidase A cerebral inhibitors** 

opposes the vasoconstrictor effect of Ang II.

caused by endogenous ouabain.

Ang III more sensitivity and a more prolonged increase in BP was observed in genetically hypertensive rats (GHR) than in normotensive Wistar-Kyoto (WKY) and Sprague-Dawley rats. But if previously treated with bestatin (an inhibitor of AlaAP and ArgAP), which prevents the conversion of Ang III to Ang IV, elevation of BP was enhanced and prolonged. These results indicate, therefore, that dysfunction in the central aminopeptidases activity could lead to Ang II and Ang III act longer and therefore, could carry out a progressive and sustained elevation of BP in RGH rats.

Later, Jensen and colleagues showed that in addition to bestatin, a GluAP inhibitor (amastatine), was injected by intracerebroventricular via, which inhibited the formation of Ang III, which is induced BP increase in rats WKYy the GHR so that there should be an effect mediated by the brain RAAS. They also noted that genetically hypertensive rats were more sensitive to the action of inhibitors than the normotensive ones. (Jensen et al., 1989)

RB150 is a prodrug with a specific inhibitory action on aminopeptidase A EC33, when administered intravenously, it inhibits cerebral aminopeptidase A, the Ang III formation and reduces BP over 24 hours in DOCA-salt rats. Thus aminopeptidase A cerebral inhibitors represent a potential antihypertensive treatment. (Bodineau et al., 2008)

In conclusion, although the discovery of ACE inhibitors and ARBs were two important milestones in the treatment of hypertension, the study of other RAAS components that act at both peripheral and central levels offer new therapeutic possibilities. The cerebral Ang III is a potent hypertensive factor. However, Ang 2-10 seems to contribute more to reduce hypertension. The results we have so far indicate that they fundamentally prevent the formation of cerebral Ang III, or perhaps also facilitates the formation of Ang 2-10, a line of research that could develop possible treatments for hypertension.

Therefore, recent studies on central inhibitors of GluAP, responsible for the formation of Ang III, may provide promising results. It is also increasingly clear that to properly understand the brain's control of the BP, studies should consider the bilaterally of the peripheral and central nervous system. The development of agonists and antagonists specific of the ACE-2, may offer an understanding of the pathophysiological role of ACE 2 in the modulation of the BP.

#### **8. Modulators of the endocannabinoid system**

The endocannabinoid system (ECS) is a new regulatory system capable of modulating a variety of physiological effects, consisting of endogenous ligands, specific receptors and mechanisms of synthesis and degradation. Endogenous ligands are a new class of lipid regulators among which there are amides and esters of polyunsaturated fatty acid chain. Endocannabinoids are defined as endogenous compounds, produced in different organs and tissues, capable of binding to cannabinoid receptors. The cannabinoids are synthesized "on demand", when they are needed, and released abroad immediately after their production. Its major molecular targets are the cannabinoid receptors (CB) type 1 and type 2 (Brown, 2007). The CB1 receptor is predominantly expressed in the central nervous system, but is also present at much lower, yet functionally relevant levels in various peripheral tissues, including the myocardium, postganglionic autonomic nerve terminals, and vascular endothelial and smooth muscle cells as well as the adipose tissue,liver, and skeletal muscle. The expression of CB2 receptors was thought to be limited to hematopoietic and immune

New Therapeutics in Hypertension 11

The U-II binds to an urotensin II receptor, Gq protein (UT), originally known as orphan GPR14 receptor. This receptor has been identified in central nervous system cells, the bone marrow, the kidneys, the heart, the vascular smooth muscle and the endothelial cells. (Ames et al., 1999) Using immunohistochemistry techniques, U-II has been found in blood vessels of the the heart, the pancreas, the kidney, the placenta, the thyroid, the adrenal glands and in the umbilical cord. UT stimulation induces the release of NO, prostacyclin, prostaglandin E2 and endothelium derived factors that balance the contractile effect on vascular smooth muscle. Vasoconstriction is mediated by receptors in the vascular smooth muscle, whereas the vasodilatation is mediated by the endothelium. However, in HF and essential hypertension, the U-II loses its vasodilator capacity. So the U-II causes vasoconstriction of

The complex and contrasting vascular actions of U-II is not only dependent on the condition of the endothelium, but also on the type of vascular bed and species. (Ames et al., 1999, Liu

In healthy individuals, the U-II acts as a chronic regulator of vascular tone. In patients with cardiovascular diseases, the balance is lost and elevated plasma levels of U-II in patients with HF, carotid atherosclerosis, kidney failure, diabetes mellitus, liver cirrhosis with portal

Endothelial dysfunction causes vasoconstriction or inadequate vasodilatation resulting in a myocardial ischemia and hypertension, associated with an increase in the U-II and UT. In fact, in patients with hypertension U-II is increased 3 or 4 times, and has shown positive correlation between HF and plasma levels of U-II. (Matsushita et al.,2001, Douglas et

Today there are several drugs that act on antagonism receptor of U-II: urantide, BIM-23 127, SB-611 812, palosuran. All at different stages of clinical research, so that in the coming years we will have results that allow us to evaluate the effectiveness on BP control and its impact

The openers of potassium channels (KCOs) are a class of drugs with an extreme chemical diversity; they include different structural classes such as benzopirans, cyanoguanidines, thioformamides, and pyrimidines. They base their action on the increase of transmembrane K+ flow, resulting in a hyperpolarization of the cell membrane through the opening of potassium channels and the closing of Ca2+ channels, as a result the cell is less excitable and less prone to stimulation. There are two large families of potassium channels described in smooth muscle: channels regulated by voltage and channels regulated by ATP. (Moreau., et al. 2000) Potassium channels, regulated by intracellular ATP are located in the heart and blood vessels and are important modulators of cardiovascular function. The opening of potassium channels produces an increase in K+ efflux from the cell so that the membrane potential at rest becomes more negative (hyperpolarization) and this leads to inhibition of calcium entry or an indirect antagonism of calcium, causing a drop in intracellular calcium concentration, relaxation of vascular smooth muscle cells and therefore vasodilatation.

endothelium independently and vasodilatation of endothelium dependently.

et al.,1999, Matsushita et al.,2001, Maguire et al., 2004, Zhu et al.,2006)

al.,2002, Richards et al.,2002, Cheung et al.,2004; Suguro et al.,2007)

on cardiovascular diseases. (Tsoukas, Kane & Giaid A., 2011)

**10. Potassium channel openers** 

hypertension and essential hypertension is found.

cells, but they have recently been identified in the brain, the liver, the myocardium, and in the human coronary endothelial and smooth muscle cells. (Pacher et al., 2008)

Increased ECS activity also contributes to the generation of cardiovascular risk factors in obesity/metabolic syndrome and diabetes such as plasma lipid alterations, abdominal obesity, hepatic steatosis, and insulin and leptin resistance. However, the ECS may also be activated as a compensatory mechanism in various forms of hypertension where it counteracts not only the increase in SP, but also the inappropriately increased cardiac contractility through the activation of CB1 receptors. In addition, the activation of CB2 receptors in endothelial and inflammatory cells by endogenous or exogenous ligands was found to limit the endothelial inflammatory response, the chemotaxis, and the adhesion of inflammatory cells to the activated endothelium with the consequent release of various proinflammatory mediators, which are key processes in the initiation and progression of atherosclerosis and reperfusion injury as well as smooth muscle proliferation. (Pacher et al., 2008)

Currently, several types of endogenous cannabinoids have been identified, all of lipid nature and derived from polyunsaturated fatty acids of long chain. Anandamide (Naraquidonoiletanolamina, AEA) and 2-arachidonoylglycerol (2-AG) are the two best known endocannabinoids. AEA may be a partial or full agonist of CB1 receptors with low action over CB2 and 2-AG an agonist both of CB1 and CB2 receptors (Bisogno., 2008, Howlett., 2002).

The possible antihypertensive effect of cannabinoid ligands is based on the lowering of BP seen in the chronic use of cannabis in humans and in response to the acute or chronic administration of tetrahydrocannabinol in experimental animals. (Bisogno., 2008)

Recently there are results that demonstrate its high hypotensive efficiency in hypertensive animals compared with the normotensive ones and evidence that the tonic activation of the ECS in various experimental models of hypertension could be a possible compensatory mechanism. (Batkai et al., 2004, Wheal et al.,2007) This hipotensive tone could be attributed to a decrease in the mediated activity by the CB1 receptor in myocardial contractility rather than vascular resistance. So preventing the degradation of endogenous AEA by pharmacological inhibition of amidohydrolase fatty acid increases myocardial levels of AEA reducing BP and cardiac contractility in hypertensive rats but not in the normotensive ones. Perhaps the amidohydrolase fatty acid could be a therapeutic target in hypertension, where its inhibition would not only reduce BP but also prevent or stop the development of cardiac hypertrophy. (Lépicier et al.,2006)

The adverse side effects of these new drugs, their interaction with other ones, their pharmacokinetics, and their efficacy in the medium and long term, their probable use in other diseases are among the many questions that remain to be clarified in this new but promising group of antihypertensive drugs.
