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A Brief History of Nitric Oxide Scientific Research

2012 Qivana

Physician Thomas Lauder Brunton first recognized the physiological effects of Nitric Oxide in 1876. Brunton experimented on a patient with angina by placing nitrite drops on a cloth for the patient to inhale. Within one minute, the patient’s pulse improved, his face became flushed with blood and his pain subsided. Brunton concluded that the nitrite compound caused blood vessels to dilate, decreasing chest pain. However, he did not know the specific molecule that caused the effect nor its mechanism of action.

In the hundred years following Brunton’s discovery, scientists further explored the effects of Nitric Oxide on the cardiovascular system. English pharmacologist Henry Dale performed several experiments on vasodilators in the early 1900s, including work with amyl nitrite, a biological source of Nitric Oxide.

Dale found that amyl nitrite operates peripherally to dilate blood vessels without acting through the nervous system. Scientists did not identify Nitric Oxide as the messenger molecule or understand its action until the late 20th century.

In the mid-1970s, Albanian-American physician Ferid Murad demonstrated that nitrite-containing compounds stimulated soluble guanylate cyclase, an enzyme found in smooth muscle that increases cyclic guanosine monophosphate (cGMP) levels. Murad demonstrated that this rise in cGMP caused smooth muscle relaxation, dilating blood vessels. The researchers proposed that this smooth muscle relaxation occurred via the action of Nitric Oxide, although they did not experimentally demonstrate this association.

Murad, Ignarro, and Robert F. Furchgott received the 1998 Nobel Prize in Physiology or Medicine identifing Nitric Oxide.

In 1981, pharmacologist Louis J. Ignarro performed the experiments that identified Nitric Oxide as a gaseous messenger molecule capable of acting on smooth muscle and other types of cells. As a result of these experiments, Murad, Ignarro, and Robert F. Furchgott received the 1998 Nobel Prize in Physiology or Medicine.

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The Swedish Nobel committee commented, “The signal transmission by a gas that is produced by one cell, penetrates through membranes and regulates the function of another cell, represents an entirely new principle for signaling in biological systems.” Before its identification as a molecule responsible for circulatory system improvement, Nitric Oxide was thought to be a waste byproduct of fossil fuel combustion and other chemical reactions.

The researchers who discovered Nitric Oxide and first explored its actions were hailed for their profound contribution to the understanding of cardiovascular health. In 1998, the then-president of the American Heart Association, Dr. Martha Hill, remarked, “the discovery of Nitric Oxide and its function is one of the most important in the history of cardiovascular medicine.” Since the Nobel Prize award in 1998, thousands of scientific articles have further explored the association between Nitric Oxide activity and cardiovascular health.

Nitric Oxide Action Mechanisms

cGMP-Mediated Pathways

Once created through NOS-dependent pathways or the nitrate/nitrite conversion pathway, Nitric Oxide acts on its physiological targets through several mechanisms. A primary mechanism of NO action is through cGMP-dependent signaling. Soluble guanylyl cyclase, or sGC, is directly activated by Nitric Oxide.

In turn, sGC activates cyclic guanosine monophosphate, or cGMP. This important signaling molecule acts upon a variety of physiological targets, including cells in the cardiovascular, nervous and digestive systems.

For example, Nitric Oxide in endothelial cells of blood vessels diffuses into the blood cell lumen to activate smooth muscle receptors. These receptors trigger a cascade of events resulting in a decrease in intracellular calcium levels.

This change in calcium causes smooth muscle relaxation, increasing blood vessel diameter and facilitating blood flow. Nitric Oxide also acts upon neurons, cells in the gastrointestinal system, and other tissue types through similar cGMP-mediated processes. item2

Nitric Oxide also works through a biological pathway independent of cGMP. A group of molecules called S-nitrosothiols (RSNOs) affect circulatory system activity and other physiological processes. These S-nitrosothiols act as carriers of Nitric Oxide, allowing it to affect other cellular systems. S-nitrosothiols activate ion channel proteins, proteolytic enzymes, gene transcription factors, and proteins involved in energy transduction.

Nitric Oxide works through S-nitrosothiol activity to regulate programmed cell death, cell signaling, inflammatory responses and vascular tone. Many of these physiological processes cause the cardiovascular benefits for which NO is best known.

Nitric Oxide Production Mechanisms

L-Arginine NO Production Pathway

The most well known NO pathway is the conversion of L-arginine to Nitric Oxide. The body naturally produces Nitric Oxide as an intermediate compound in the conversion of L-arginine, a semi-essential amino acid, to nitrite. L-arginine is commonly found in dietary sources of proteins including dairy products, beef, pork, poultry, nuts, and seeds. A class of enzymes called Nitric Oxide synthases (NOS) act on L-arginine, oxidizing one of its nitrogen molecules.

The activity of NOS relies on the presence of six cofactors. Two types of NOS, endothelial NOS-3 and neuronal NOS-1, depend on calcium to oxidize L-arginine and generate Nitric Oxide. These enzyme isoforms produce relatively low levels of NO, allowing the molecule to diffuse to nearby cells to exert its effects. The other NOS isoform, inducible NOS-2, creates Nitric Oxide through a calcium-independent process. NOS-2 produces larger amounts of Nitric Oxide than the calcium-dependent enzyme isoforms.

The NO that is formed through the L-arginine reaction exists as a gaseous molecule that lasts only a few seconds. Nitric Oxide is highly reactive, meaning that it easily reacts with other molecules to form new compounds. The most prominent example of Nitric Oxide reactivity is the reaction of NO with oxygen to form nitrogen dioxide.

As a result of its reactivity, Nitric Oxide has the ability to act upon nearby cells but cannot travel long distances to act upon faraway physiological targets. This makes NO ideally suited to act as a messenger with localized action between groups of related cells.

Many products on the market are entirely based on the conversion of L-arginine. An entire category of bodybuilding supplements is being marketed as Nitric Oxide producing, all using this pathway.

However L-arginine supplements have significant limitations. Because this pathway is highly complex and has 6 required co-factors, very little L-arginine is actually converted into NO. In fact there are at least 8 different pathways L-arginine can be used in, other than Nitric Oxide, so less than 3% of L-arginine is actually converted into NO.

Therefore, supplementation with L-arginine has serious limitations and studies show it can make many health conditions worse if it goes unconverted. This is especially true in patients with existing heart conditions and in people over the age of 60.

In one notable study the researchers in the Journal of the American Medical Association concluded that L-arginine will probably only work in people with “preexisting L-arginine deficiency” (such as chronic kidney disease or people with elevated ADMA). They also noted that people over age 60 wouldn’t experience any benefit as they undergo metabolic changes that make inhibit their use of the amino acid.

Finally, they noted that without NOS cofactors guaranteeing production of NO, L-arginine generates cell damaging “reactive oxygen species.”

So while L-arginine can stimulate production of NO and provide the benefits of NO, supplements based entirely on L-arginine are limited in their efficacy and safety profile.

Alternate NO Production Pathways

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In addition to Nitric Oxide production from the conversion of L-arginine, the body also creates NO through a pathway independent of NOS activity. People receive nitrate and nitrite through dietary sources such as green leafy vegetables, certain root vegetables, meat products, and drinking water. Several strains of anaerobic bacteria in the tongue and in the gastrointestinal tract perform a reduction reaction that converts nitrate to nitrite. Human cells cannot effectively metabolize nitrate, meaning that the body relies on oral bacteria to perform this nitrate reduction function.

Approximately 25 percent of nitrate from dietary sources is secreted in saliva. Of that salivary nitrate, nearly 20 percent is converted to nitrite by these oral probiotic bacterial strains. The result of this bacterial activity is a concentration of nitrite in the saliva; salivary nitrite levels are nearly 1,000 times higher than plasma nitrite levels.

As the saliva travels through the digestive system, it comes into contact with gastric acid in the stomach. Approximately 1 to 1.5 liters of saliva enter the stomach in this way every day. When nitrite comes into contact with stomach acid, it reacts to form nitrous acid, or HNO2.

The gastric acid further reduces nitrous acid to form NO. This reaction occurs independently of NOS, constituting a separate pathway of Nitric Oxide production. This NO synthesis pathway depends on dietary intake of nitrate and nitrite, not the presence of L-arginine.

Although considerable Nitric Oxide production occurs in the reaction of nitrite with gastric acid, not all-available nitrite is converted in this way. The gastrointestinal system absorbs excess nitrite, storing it in cells throughout the body. Once within the cells, nitrite can form Nitric Oxide through a reaction with a ferrous heme protein. However, this process does not produce NO as efficiently as other pathways. The storage of nitrite within cells creates a reserve pool that the body can access to create more NO molecules.

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"The discovery of Nitric Oxide is one of the most important in the history of cardiovascular medicine" ~ President of the American Heart Association
 


"Nitric Oxide is the body's way of protecting against cardiovascular disease" ~ UCLA Department of Molecular and Medical Pharmacology
 


"Any risk factor for cardiovascular disease is associated with loss of Nitric Oxide." ~ Mayo Clinic Research

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