Dramatic increase of nitrite levels in hearts of anoxia-exposed crucian carp supporting a role in cardioprotection

Abstract

Sandvik GK, Nilsson GE, Jensen FB. Dramatic increase of nitrite levels in hearts of anoxia-exposed crucian carp supporting a role in cardioprotection. Am J Physiol Regul Integr Comp Physiol 302: R468 -R477, 2012. First published November 30, 2011 doi:10.1152/ajpregu.00538.2011 Ϫ ) functions as an important nitric oxide (NO) donor under hypoxic conditions. Both nitrite and NO have been found to protect the mammalian heart and other tissues against ischemia (anoxia)-reoxygenation injury by interacting with mitochondrial electron transport complexes and limiting the generation of reactive oxygen species upon reoxygenation. The crucian carp naturally survives extended periods without oxygen in an active state, which has made it a model for studying how evolution has solved the problems of anoxic survival. We investigated the role of nitrite and NO in the anoxia tolerance of this fish by measuring NO metabolites in normoxic, anoxic, and reoxygenated crucian carp. We also cloned and sequenced crucian carp NO synthase variants and quantified their mRNA levels in several tissues in normoxia and anoxia. Despite falling levels of blood plasma nitrite, the crucian carp showed massive increases in nitrite, S-nitrosothiols (SNO), and ironnitrosyl (FeNO) compounds in anoxic heart tissue. NO 2 Ϫ levels were maintained in anoxic brain, liver, and gill tissues, whereas SNO and FeNO increased in a tissue-specific manner. Reoxygenation reestablished normoxic values. We conclude that NO 2 Ϫ is shifted into the tissues where it acts as NO donor during anoxia, inducing cytoprotection under anoxia/reoxygenation. This can be especially important in the crucian carp heart, which maintains output in anoxia. NO 2 Ϫ is currently tested as a therapeutic drug against reperfusion damage of ischemic hearts, and the present study provides evolutionary precedent for such an approach. nitric oxide; S-nitrosothiols; anoxia/reoxygenation; cytoprotection; nitric oxide synthase NITRIC OXIDE (NO) is an important regulator of many biological functions in vertebrates. In normoxic conditions, NO is produced from L-arginine and O 2 by NO synthases (NOS). The classical role is in vasodilation, where NO binds to the heme of the guanylyl cyclase in smooth muscle cells, leading to a rise in cGMP and subsequent muscle relaxation (35). NO also binds to heme groups of other proteins and is involved in reactions with thiols and secondary amines of different proteins, leading to formation of iron-nitrosyls (FeNO), S-nitrosothiols (SNO), and N-nitrosamines (NNO), respectively (26). Through formation of FeNO and SNO, NO can activate and inactivate enzymes or modify protein function (17,73). NO is inactivated by oxidation to nitrate (NO 3 Ϫ ) in reaction with oxygenated hemoglobin and myoglobin, and it is metabolized to nitrite (NO 2 Ϫ ) in reaction with O 2 (48). Nitrite was previously considered an inert end product of this oxidation, but nitrite can be reduced back to NO, and it has become clear that nitrite functions as a NO storage pool that is activated by va...

Globin X is a six-coordinate globin that reduces nitrite to nitric oxide in fish red blood cells

“…In this regard, it is now recognized that deoxygenated Hb in mammalian blood can reduce nitrite to NO in vivo, and thus modulate blood pressure, hypoxic vasodilation, mitochondrial respiration and the inhibition of platelet activation (11,35). The function of NO and nitrite in fish metabolism has been explored only recently (36)(37)(38). Nitrite and nitrate levels detected in the bass plasma are in similar ranges to the reported values for carp and goldfish (Fig.…”

Circulating nitric oxide metabolites and cardiovascular changes in the turtleTrachemys scriptaduring normoxia, anoxia and reoxygenation

“…Previous studies on humans and rats have also ascribed the formation of FeNO in red blood cells to the nitrite reductase activity of deoxy-Hb during acute hypoxia (Cosby et al, 2003;Feelisch et al, 2008). In other studies on goldfish and crucian carp, plasma nitrite decreases while erythrocyte nitrite is unchanged or increased during days of exposure to hypoxia or anoxia (Hansen and Jensen, 2010;Sandvik et al, 2012). In contrast to these studies, however, nitrite did not change significantly in plasma or red blood cells after the turtles had been exposed to 3h of anoxia (Fig.3).…”

“…In turtle, normoxic red blood cells contained high levels of nitrite (1.1moll -1 ) as in crucian carp (1.4moll -1 ) (Sandvik et al, 2012) (Table1). Normoxic red blood cell values of RSNO (0.2moll -1 ) were in between those of humans and goldfish (~0.05moll -1 ) (Hansen and Jensen, 2010;MacArthur et al, 2007) and those of crucian carp (~1moll -1 ) (Sandvik et al, 2012) and Tibetan human populations (2.4moll -1 ) (Erzurum et al, 2007). Notably, while being present at very low (often undetectable) concentrations in other species, FeNO+RNNO complexes in red blood cells were present in appreciable amounts (1.9moll -1 ) in Trachemys turtles under normoxic conditions (Table1), indicating that the fate of NO is different in different species.…”

“…In the turtle, it is possible that nitrite for erythrocyte NO generation might have originated from some of the tissues that were perfused by blood during anoxia. It is known that nitrite permeates cellular membranes and is transported into red cells and tissues upon an elevation of extracellular levels (Bryan et al, 2005;Feelisch et al, 2008;Jensen and Rohde, 2010) or exposure to reduced O 2 levels (Hansen and Jensen, 2010;Sandvik et al, 2012). Measuring nitrite in tissues other than blood in future studies will help to establish the role of nitrite as a source of NO during anoxia in turtles.…”

Metabolic fates and effects of nitrite in brown trout under normoxic and hypoxic conditions: blood and tissue nitrite metabolism and interactions with branchialNOS,Na+/K+-ATPaseandhsp70expression

“…Nitrite is generated endogenously as an oxidative metabolite of the signaling molecule nitric oxide (NO) produced by nitric oxide synthases (NOS), giving rise to nitrite concentrations at or below the micromolar range in blood and tissues of fish (Hansen and Jensen, 2010;Sandvik et al, 2012) and mammals (Bryan et al, 2005). The natural pool of nitrite functions as a reservoir of NO availability, from which NO can be regenerated under hypoxic conditions via a number of cellular proteins (including deoxygenated hemoglobin and myoglobin) that reduce nitrite to NO, allowing a continued NO production when NOS enzymes become compromised by a lack of O 2 (Lundberg et al, 2008;Jensen, 2009).…”