The Relationship between Estrogen and Nitric Oxide in the Prevention of Cardiac and Vascular Anomalies in the Developing Zebrafish (Danio Rerio)

Abstract

It has been known that both estrogen (E2) and nitric oxide (NO) are critical for proper cardiovascular system (CVS) function. It has also been demonstrated that E2 acts as an upstream effector in the nitric oxide (NO) pathway. Results from this study indicate that the use of a nitric oxide synthase (NOS) inhibitor (NOSI) which targets specifically neuronal NOS (nNOS or NOS1), proadifen hydrochloride, caused a significant depression of fish heart rates (HR) accompanied by increased arrhythmic behavior. However, none of these phenotypes were evident with either the inhibition of endothelial NOS (eNOS) or inducible NOS (iNOS) isoforms. These cardiac arrhythmias could also be mimicked by inhibition of E2 synthesis with the aromatase inhibitor (AI), 4-OH-A, in a manner similar to that of nNOSI. In both scenarios, by using an NO donor (DETA-NO) in either NO + nNOSI or E2 + AI co-treatments, fish could be significantly rescued from decreased HR and increased arrhythmias. However, the addition of an NOS inhibitor (L-NAME) to the E2 + AI co-treatment fish prevented the rescue of low heart rates and arrhythmias, which strongly implicates the NO pathway as a downstream E2 targeted molecule for the maintenance of healthy cardiomyocyte contractile conditions in the developing zebrafish. Cardiac arrhythmias could be mimicked by the S-nitrosylation pathway inhibitor DTT (1,4-dithiothreitol) but not by ODQ (1H-[1–3]oxadiazolo[4,3-a]quinoxalin-1-one), the inhibitor of the NO receptor molecule sGC in the cGMP-dependent pathway. In both the nNOSI and AI-induced arrhythmic conditions, 100% of the fish expressed the phenotype, but could be rapidly rescued with maximum survival by a washout with dantrolene, a ryanodine Ca2+ channel receptor blocker, compared to the time it took for rescue using a control salt solution. In addition, of the three NOS isoforms, eNOS was the one most implicated in the maintenance of an intact developing fish vascular system. In conclusion, results from this study have shown that nNOS is the prominent isoform that is responsible, in part, for maintaining normal heart rates and prevention of arrhythmias in the developing zebrafish heart failure model. These phenomena are related to the upstream stimulatory regulation by E2. On the other hand, eNOS has a minimal effect and iNOS has little to no influence on this phenomenon. Data also suggests that nNOS acts on the zebrafish cardiomyocytes through the S-nitrosylation pathway to influence the SR ryanidine Ca2+ channels in the excitation-coupling phenomena. In contrast, eNOS is the prominent isoform that influences blood vessel development in this model.

Sex Disparities in Sudden Cardiac Death

“…[32][33][34][35][36][37] Less is known about the role of estrogen in SCD. Studies investigating estrogen's role in the development of ventricular arrhythmias have found both proarrhythmic and antiarrhythmic effects in females or both sexes, [38][39][40][41][42][43] indicating a need for further investigation.…”

Inhibition of cytochrome P450 with proadifen alters the excitability of brain catecholamine-secreting neurons

“…The main limitations of this study are the use of a nonselective CYP inhibitor and non-distinguishing between brain and hepatic CYP inhibition. The involvement of the inhibitory effect of SKF525 on the nitric oxide synthase (NOS; Sykes et al 2016) cannot be excluded as well. In future studies, the effect of the selective inhibitors of the specific CYP subtypes, such as CYP3A1, CYP3A2, CYP3A4, and CYP3A5, which are fundamental in glucocorticoid metabolism (Peng et al 2011), should be tested.…”

Cardiac Hypoxia Tolerance in Fish: From Functional Responses to Cell Signals

“…Data on trout alevins ( Salmo trutta ) show that L-arginine-derived NO reduces heart rate, while NOS inhibition induces tachycardia, an event that was considered an indirect consequence of vasoconstriction [ 119 ]. However, a direct chronotropic control mediated by the gas is reported in developing zebrafish, in which NOS inhibition was found to depress heart rate, and also to induce arrhythmic behavior [ 120 ]. Interestingly, in fish NO is involved also in shaping early cardiac development.…”