Nitric Oxide

Biological Significance

Structure

Nitric Oxide (henceforth NO - depicted below) is a small signalling molecule synthesized from the amino acid L-arginine by the family of nitric oxide synthases including eNOS (endothelial, NOS-III), iNOS (inducible, NOS-II), and nNOS (neuronal, NOS-I). This family of enzymes work as dimers, along with multiple co-factors including tetrahydrobiopterin, flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN), iron, and zinc. While the regulation and modulation of each isoform differs significantly, all of the isoforms catalyze the reaction of L-arginine with NADPH and oxygen, to yield NO, citrulline and NADP (Knowles and Moncada (1994); Marletta (1994).

How NO Signals

The elucidation of nitric oxide as a signaling gas molecule resulted in the award of the Nobel prize in physiology/medicine in 1998, as this was the first time a gas molecule was identified to be produced by one cell, readily diffusing to another cell, and then acting as a signaling molecule in it. For example, NO produced by eNOS in endothelial cells diffuses to the adjacent smooth muscle cells, where it initiates a cascade of reactions by activating the soluble guanlyate cyclase, which catalyzes the production of cyclic GMP ^[1]. Elevation of cGMP leads to the activation of protein kinase G (PKG), which in turn phosphorylates myosin light chain (MLC) phosphatase (thus activating it). In turn, activated MLC phosphatase dephosphorylates MLC, resulting in relaxation of smooth muscle cells, and thus, vascular relaxation.

Nitric oxide signals by stimulating its receptor, the soluble guanylyl cyclase receptor, and increasing cellular levels of the signalling molecule called cyclic guanidine monophosphate (cGMP)

Additional players in the regulation of vascular tone include the phosphodiesterase family (PDE 1 – 11), which catalyze the hydrolysis of cGMP at the 3’ end ^[2][3], effectively stopping NO-mediated vascular relaxation. Due to the tight regulation of eNOS and NO production, it is difficult to modulate vascular relaxation by influencing eNOS activity.

Due to the physiologic relevance of PDEs in controlling cGMP levels, these have become a popular target when vascular relaxation and blood flow are desired. An example of this include drugs such as Viagra, Cialis and Levitra, all of which inhibit PDE-5, which happens to be specifically expressed in the smooth muscle cells within the corpus cavernosum in the penis ^[4]. Since inhibition of these enzymes result in accumulation of cGMP, it essentially becomes possible to potentiate the vascular relaxation effects of NO.

Phosphodiesterases are negative regulators of cGMP and cAMP (they hydrolyze these molecules). While not all PDE enzymes can target cGMP induced by NO effects on guanylate cyclase, a few of them have the ability to control NO signaling via degradation of key intermediate signaling molecule (cGMP)

Oxidative Potential

NO can potentially be degraded into a molecule known as peroxynitrate (OONO-) which is the result of NO reacting with superoxide anions (O₂^-).^[5][6] OONO- also acts as a reactive signalling molecule, although the end results tend to be formation of several structures known to be negative to the body; OONO- can nitrosylate (donate a nitrogen group) to amino acids to form compounds such as 3-nitrotyrosine or S-nitrosocysteine,^[7] formation of protein carbonyls, or nitrosylation of phospholipids containing polyunsaturated fatty acids (PUFAs).^[8][9] In this sense, Nitric Oxide can be used as a substrate by superoxide to form reactive compounds that are negative to health despite NO being relatively benign.^[8]

Nitric Oxide can be transformed (by combining with superoxide radicals) to form peroxynitrate, which then can produce a variety of molecules that are associated with a state of unhealth and thought to play a role in the pathology

Pharmacology

Supplemental Nitric Oxide

NO that is synthesized in the body and subsequently released into the blood tends to have a half-life of 5 seconds or less,^[10][11] and in vitro some complexes can be made to prolong the half-life to 445s or so for research purposes.^[12] These short half-lifes indicate quick degradation of the Nitric Oxide molecule into its components (Nitrogen and Oxygen), and propert storage of NO can prolong its shelf-life has only been confirmed for up to 5 days^[13] using Mylar balloons which attenuate degradation.^[14] Due to this poor stability ex vivo (outside of the body), Nitric Oxide per se is never used as a supplement; but rather compounds that can stay in the blood for long enough to continuatelly aid in the production of new NO.

Nitric Oxide, per se, is unstable and has a short half-life; it exerts its benefits rapidly, but has no role as a supplment in and of itself. NO supplementation required other compounds that influence the internal NO production system

Physiology

Cardiovascular

Nitric Oxide is involved in relaxation of vascular smooth muscle, which is a mechanism underlying cardioprotective effects from Nitric Oxide (via reducing blood pressure)^[15]

Neuronal

Nitric Oxide tends to modulate ion channels, intrinsic excitability, mediates synaptic plasticity, and can diffuse through cellular membranes.^[16]

Neuronal nitric oxide synthase (nNOS) is able to form a dimer with a protein known as PSD95^[17][18] and this complex appears to be a positive regulator of depression, as inhibiting the nNOS-PSD95 interaction has antidepressant effects.^[19] This complex is activated downstream of NMDA receptor activation.^[20]

Supplementation

Nitric Oxide Donors

Some supplements aid NO production merely be being a source of Nitrogen that the NOS enzyme can use to produce NO with. Arginine is the standard NO donor in supplementation, and Citrulline is a more bioavailable form of Arginine. Other NO donors that exist are S-Nitrosoglutathione (formed endogenously) or the two classes of N-diazeniumdiolates or S-nitrosothiols, the latter of which contains the endogenous S-Nitrosoglutathione.^[21][22]

Some compounds merely provide nitrogen for the enzyme to use to produce NO