Nitric Oxide and Nitric Oxide Synthase, Biology, Pathology, Localization.Pdf
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Hislol Hislopalhol (1997) 12: 251-261 Histology and 001: 10.14670/HH-12.251 Histopathology http://www.hh.um.es From Cell Biology to Tissue Engineering Invited Review Nitric oxide and nitric oxide synthase: biology, pathology, localization G. Wolf Institute of Medical Neurobiology, University of Magdeburg, Magdeburg, Germany Summary. Nitric oxide (NO) has opened a new and affected tissues (Knowles and Moncada, 1994; Moncada vigorous field of biological and clinical experimentation et aI., 1994a,b; Schmidt and Walter, 1994; Body et aI., as evidenced presently by about one hundred original 1995; Gross and Wolin, 1995; Kerwin et aI., 1995; publications every week. Being a biological signal under Moncada and Higgs, 1995; Schroeder and Kuo, 1995; physiological conditions, NO may be «foe or friend» to Vincent, 1995). pathologically affected tissues. Major insights into the biology and pathology of this unorthodox biomolecule Nitric oxide, an unorthodox biomolecule have come from the histochemical analysis of NO synthase (NOS) and its molecular isoforms that are NO is a gaseous radical produced in the atmosphere responsible for the formation of NO. Immunocyto by lightning and the burning of fossil fuels. Since the chemistry as well as NADPH-diaphorase histochemistry late 1970s NO is known to activate guanylyl cyclase and are most widely used to visualize NOS in various to cause vascular smooth muscle relaxation (Arnold et tissues. There are several constraints regarding aI., 1977; Gruetter et aI., 1979). But it took several years specificity and sensitivity of the techniques used and, to discover that inorganic oxides of nitrogen are therefore, apparent discrepancies in the literature biologically produced (Green et aI., 1981) and that NO is concerning the cellular and subcellular distribution of identical to the so-called "endothelium derived relaxing NOS and its isoforms. Despite such inconsistencies there factor" (EDRF) (Furchgott, 1988; Ignarro et aI., 1988). is a wealth of data that has an important impact on Since that time there has been, despite initially further investigation of NO-mediated processes considerable scepticism, an explosion of research underlying a vast number of NO-mediated physiological activity on this topic. Named the molecule of the year in and pathological mechanisms. 1992, NO has opened a new and vigorous field of biological and clinical experimentation as evidenced Key words: Nitric oxide, Nitric oxide synthase, presently by nearly one hundred original publications Histochemistry, Electron microscopy every week. Due to its unique ability to diffuse quickly in both aqueous and lipid environment, NO is assumed to spread Introduction rapidly to neighbouring tissue elements regardless of intervening membranes. Derived from theoretical Nitric oxide (NO) has recently expanded its status considerations, Wood and Garthwaite (1994) have from an environmental pollutant to a biological signal calculated that the physiological sphere of influence of a molecule that mediates blood vessel relaxation and single point source of NO emitting for 1-10 sec has a immune responses , kills pathogens, inhibits platelet diameter of about 200 JI m. Contrary to conventional aggregation and adhesion, and serves as a neuro biosignals that act via specific receptor molecules, NO modulator in the central and peripheral nervous system. functions in a fairly specific manner by a wide range of There is, moreover, evidence that NO may be a chemical reactions controlling enzyme activities, ion physiological regulator of mitochondrial respiration and channels, gene transcription, mitochondrial respiration, gene activity, and that NO is implicated in a variety of and can interact with oxygen-derived radicals to produce human diseases being, in dependence on the particular other highly reactive substances (Brown, 1995; Crow circumstances, a «foe or friend» to pathologically and Beckman, 1995; Garthwaite and Boulton, 1995; Gross and Wolin, 1995; Moncada and Higgs, 1995). Offprint requests to: Prof. Dr. Gerald Wolf, Institute of Medical There are compelling reasons to believe that NO Neurobiology, Otto·von-Guericke-University, Leipziger Str. 44, 0-39120 mediates a variety of functions in the nervous system, Magdeburg, Germany sLlch as synaptic transmission, plasticity, regulation of 252 Nitric oxide and nitric oxide synthase cerebral blood flow, induction and regulation of the underscoring the importance of large-scale experiments circadian rhythm, and hyperalgesia (for review, see on the NO-biology as well as clinical trials to develop Schuman and Madison , 1994; Garthwaite and Boulton, the NO pharmacology and to assess undesirable side 1995; Paakari and Lind berg , 1995; Bachneff, 1996). effects that might arise from pharmacotherapeutic Furthermore, NO is involved jn the development of interventions (Body et aL , 1995; Moncada and Higgs, tolerance to and withdrawal from morphine (Bhargava, 1995; Schroeder and Kuo, 1995). 1995; Vaupel et aL , 1995) and alcohol (Lancaster, 1995). At higher concentrations NO has been proposed to have Nitric oxide synthase antibacterial , antifungal , anti protozoal and tumoricidal activity (Vallance and Collier, 1994; Kerwin et aL, 1995; Major insights into the biology and pathology of NO Schoedon et aI. , 1995). NO overproduction in have come from the characterization of the enzyme that mammalian systems may contribute to cell damage or is responsible for its formation in living tissues, called cell death (Dawson, 1995; Gross and Wolin, 1995; NO synthase (the term «synthase» stands, in contrast to Krbnke et aI., 1995) (Fig. I). Thus , NO might be «synthases» , for enzymes that do not utilize ATP). implicated in cerebral disorders induced by hypoxia and Catalyzed by NO synthase (NOS; EC 1.14.13.39), NO is ischemia, migraine, Parkinson 's disease and AIDS produced in a NADPH-dependent manner by oxidation dementia (Moncada, 1994; Olesen et aL , 1995). On the of the guanidino nitrogen of L-arginine in the presence other hand, manipulation of the NO pathway may offer of molecular oxygen and several cofactors (FMN, FAD , therapeutic benefit in selected human diseases tetrahydrobiopterin , heme moiety). Three major Argi· 02 C . 15 NOS .... nine~NNOS ::(P " \ \ Citrul \ line \ \ Glu \ "-. Guanylyl cyclase GTP --+ cGMP l various f cellular functions • NO· Ca++- Calmodulin FMN , FAD NOS H4-Blopterin Fig. 1. Scheme of NO effects at a CNS glutamate synapse, partially hypothetical. Synaptic activity results in an increase in pre· and postsynaptic Ca++ . levels via voltage· or transmitter(NMDA·receptor)·gated ion channels (a). Calcium binds to calmodulin and activates (b) nitric oxide synthase (NOS, neuronal isoform). NO generated by the conversion of arginine to citrulline (c) modulates synaptic activity (d) , dilates blood vessels (e) and influences many other cellular functions by modification of cellular proteins, e. g. guanylyl cyclase (f). NO also reacts with superoxi de anions (0 2 ' ) to peroxynitrite (ONOO') that may, in the case of a pathological overproduction, damage cells in the vicinity, possibly by initiating a cytotoxic cascade (g). 253 Nitric oxide and nitric oxide synthase isoforms of NOS have been identified so far. Based on synthesizing enzyme NOS. the historical order of purification and cDNA isolation, Several topochemical methods, above all immuno or on the cell type or organ in which the enzyme was cytochemistry, NADPH-diaphorase histochemistry and originally discovered, the isoforms are termed: NOS I in-situ hybridization of NOS-mRNA, can be used to (brain or neuronal NOS, nNOS), NOS II (macrophage or demonstrate NOS expression under in-situ conditions hepatocyte NOS), and NOS III (endothe li a l NOS , (Vincent, 1994; Beesley, 1995) . Moreover, there are a eNOS) (Forstermann and Kleinert , 1995; Griffith and few reports in which supplementary methods have been Stuehr, 1995). Whereas NOS r and NOS III were applied, such as the autoradiographic localization of the originally identified as constitutive in brain and vascular enzyme by demonstrating the binding of the irreversible endothelium, type II is norma lly not expressed (or NOS inhibitor [3H]L-nitroarginine to the enzyme occurs at a very low level), but is inducible by cytokines molecule (Burazin and Gundloch, 1995), and, further and bacterial products (therefore also termed «inducible more , sin g le cell PCR of NOS-mRNA , by which the NOS», iNOS). After induction, there are apparently no expression of nNOS was shown in single hippocampal regulatory mechanisms for NOS Jr . The activity of NOS neurons (Chiang et aI., 1994). r a nd III , on the other hand , is dependent on the availability of free calcium ions and calmodulin. Immunocytochemistry Molecular cloning of the constitutive NOS isoforms has indicated a consensus sequence for phosphorylation Several antisera and monoclonal antibodies have ites, which can be modulated by prote in kinase A, been raised against the different NOS-isoform proteins protein kinase C and Ca++/calmodulin-dependent protein by individual researchers, or have become commercially kinase as well (Nakane et aI., 1991 ; Dinerman e t avai lable, amongst others from Affiniti Bioreagents a L. , 1994a). The functional s ignificance of NOS (UK) , Transduction Laboratories (USA), Euro phosphorylation, showing in vitro an inhibitory effect on diag nostica (Sweden), Biomol (Germany) , Alexis the enzyme activity, is not yet clear. Corporation, and Auspep (Australia). A critical factor in As indicated by cloning studies,