Medical brain Notes 3

  Pharmacology of nitric oxide (NO)   Nitric oxide is a mediator that is very different from any other hormones and transmitters. Three key properties of NO are important to its unique mode of signal transmis-sion:   •       NO is a very small molecule and permeates cell mem-branes with ease – its membrane permeability is compa-rable to that of oxygen.   •        It binds very fast and avidly to heme, as both O 2  and CO do as well. Its affinity for heme is higher than that of O 2  but lower than that of CO. Binding of NO to heme is at the heart of its major established signalling mech-anism.   •        NO is a radical (·N=O) and therefore quite reactive. It can react with molecular oxygen and various reactive oxygen species. The ensuing products in turn may react with amino acid side chains in proteins, leading to S-ni-trosylation of cysteines and O-nitrosylation of t...

  Vascular effects of nitric oxide The pharmacological activity of nitric oxide was recog-nized before it was identified as a physiological mediator it-self. This discovery was made during an investigation into the nature of the so-called `endothelium-derived relaxing factor' (EDRF). The activity of EDRF can be triggered by the application of acetylcholine to the aorta of experimen-tal animals. In the aorta (as well as other blood vessels), not only the smooth muscle itself but also the endothelium is supplied with cholinergic nerve terminals and accord-ingly possesses acetylcholine receptors, which are of the muscarinic type (Figure 11.1a). After cutting the aorta into strips (Figure 11.1b), the endothelium can be removed me-chanically or enzymatically 1 . Aortic strips with or without endothelium will both respond with contraction to nora-drenaline or  α -selective adrenergic agonists. However, if acetylcholine is applied subsequently, only the strip that re-tains its endoth...

  Nitric oxide synthase and its isoforms NO is generated in vivo by nitric oxide synthase (NOS). This enzyme is located in the cytosol and utilizes argi-nine, molecular oxygen, and NADPH as substrates (Figure 11.4a). NOS is a fairly complex molecule that possesses multiple redox coenzymes which constitute a little electron transport chain of their own. NOS is a dimer, and the elec-tron transfer actually occurs between the two subunits (Fig-ure 11.4b). There are several subtypes of NOS: Endothelial NOS or eNOS, neuronal NOS or nNOS, and inducible NOS or iNOS. eNOS is responsible for the blood vessel-relaxing effect discussed above. Of note, it is found in both arter-ies and veins. Accordingly, its activation will both lower resistance (by arterial relaxation) and increase volume ca-pacity (by venous relaxation), and therefore cause a strong reduction of blood pressure. In fact, NO-releasing drugs are the most powerful vasodilatators available, overriding the action of other mediator...

  Biochemical mechanisms of NO signaling How does NO signalling work? As mentioned above, NO is generated within the cytosol of the endothelial cell (or, in the CNS, the presynaptic cell). As it is able to cross cell membranes with ease, it will diffuse into neighbour-ing cells, i.e the smooth muscle cells (in the blood ves-sel walls) or the post-synaptic nerve cells (in the case of nNOS). There, it will bind to a heme group that is attached to the enzyme called `soluble guanylate cyclase' (sGC). NO binding will activate sGC, which will result in the syn-thesis of cyclic guanosine monophosphate (cGMP) from GTP, in a manner analogous to adenylate cyclase, which as we've seen forms cAMP from ATP. However, the molec-ular mechanism of sGC activation by NO is quite special: Binding of NO to one side of the heme moiety of sGC will break the bond of the heme iron to a histidine residue on the opposite side, which in turn triggers the conformation-al change that leads to activation (Fi...

  Role of NO in macrophages S- and O-nitrosylation are, however, very likely important in the second function of NO – i.e., in killing microbes by macrophages (Figure 11.11). Macrophages are the most potent phagocytic cells in the immune system, in charge of dealing with hardy microbes such as mycobacteria, which are completely resistant to other phagocytic cells such as granulocytes. In these cells, the NO concentrations are sub-stantially higher than in nerve or endothelial cells. While reactive oxygen species have a substantial bactericidal ef-fect in the absence 5  of NO, the latter enhances the ability of macrophages to kill bacteria. This may partially be due again to the ability of NO to cross membranes with ease, which would let it penetrate the interior of the microbial cell; most other effector molecules (including, e.g., they very toxic superoxide anion) cannot do this. Inside the mi-crobial cell NO might again act by binding to heme, e.g. within the microbial respi...

  NO releasing drugs So, what does a NO releasing drug look like? The first one there was (Figure 11.12, top) is more widely known in an-other application, which helped Mr. Alfred Nobel make his fortune – and still maintain his good conscience, since he believed that this weapon would be so horrible that mankind would henceforth abstain from warfare (a hope now held by many for nuclear weapons). The effect of ni-troglycerine was initially noted in the form of the `Mon-day headache': The factory workers, returning to work on Monday, experienced a strong headache 8  that faded away with continuous exposure to nitroglycerine vapours dur-ing the workweek, only to reappear the next Monday after withdrawal during the weekend. Nitroglycerin – applied sublingually as a spray for rapid uptake and avoidance of liver first-pass effect – is still the standard treatment of acute events of angina pectoris, which is basically an acute, painful deterioration of coronary artery perfusion, caus...

  NOS inhibitors Inhibitors of NOS, while widely used in experimental re-search, are still in under investigation for clinical application. As mentioned above, the release of endogenous NO may be pathologically enhanced in septicaemia, leading to septic shock. Treatment with NOS inhibitors has been sug-gested as early as ten years ago but is still in the experimen-tal stage. Inhibitors of iNOS are also of interest (and at the stage of animal experiments) in the treatment of chronic in-flammatory diseases, e.g. rheumatoid arthritis. Some such drugs are derivatives of arginine (Figure 11.13a). Interest-ingly, very simple alkyl derivatives of isothiourea are very potent inhibitors of NOS (Figure 11.13b). For clinical use it would, of course, be very favourable to have isoform-se-lective inhibitors. Some experimental inhibitors that indeed do show some preference for iNOS and nNOS, respectively, are shown in Figure 11.13c. In particular, selective inhibi-tion of iNOS should be advantag...