Administration of 3‑hydroxybutyrate (3‑OHB) to animals and humans has been associated with increased cardiac output, but the mechanism remained unclear. Specifically, it was not known whether observed hemodynamic benefits reflect a direct myocardial inotropic action, or whether they result from systemic effects such as peripheral vasodilation and consequent cardiac unloading. It was also uncertain whether any myocardial effect is mediated via receptor‑dependent signaling (for example, beta‑adrenergic pathways and downstream cAMP) or through improvements in myocardial bioenergetics.
The study reported here aimed to resolve these questions by combining integrated, in vivo hemodynamic assessment of left ventricular (LV) function using pressure–volume analysis with ex vivo experiments on isolated rat LV trabeculae under controlled pharmacologic conditions.
Subjects were normotensive male HanSD rats (age 20–30 weeks). Two complementary experimental approaches were used:
In vivo left ventricular pressure–volume analysis to assess global LV contractility, ventriculo‑arterial coupling, and mechanical efficiency after administration of 3‑OHB.
Ex vivo superfused isolated left ventricular trabeculae prepared from rat hearts, electrically paced at 1 Hz and bathed in Tyrode solution, to measure twitch force responses to 3‑OHB and to probe signaling mechanisms.
In the trabeculae experiments, 3‑OHB was applied at 1 mmol/L. Pharmacologic probes included the beta‑1 adrenergic blocker metoprolol, an adenylyl cyclase inhibitor (2,5‑dideoxyadenosine), and the β‑adrenergic agonist isoprenaline, used to test whether the 3‑OHB inotropic effect required beta‑adrenergic receptor activation or cAMP‑dependent signaling pathways.
Pressure–volume analysis of the left ventricle demonstrated changes consistent with enhanced intrinsic LV contractility after administration of 3‑OHB. Key reported effects included:
An increase in left ventricular end‑systolic elastance (Ees) by approximately 34%, a load‑independent index of contractility.
Improved ventriculo‑arterial coupling by about 65%, indicating a more favorable relationship between LV contractile state and arterial load.
Increased left ventricular mechanical efficiency by roughly 28%.
Taken together, these integrated hemodynamic changes were interpreted by the authors as evidence that 3‑OHB produces a direct positive inotropic effect rather than merely reducing afterload via systemic vasodilation.
In isolated LV trabeculae experiments, 1 mmol/L 3‑OHB increased twitch force in preparations from both healthy and heart failure (HF) groups. The magnitude of force augmentation differed between groups:
Healthy hearts: mean increase in twitch force of +55.8%.
Heart failure group: mean increase in twitch force of +35.6%.
The difference in the degree of augmentation between healthy and HF trabeculae reached statistical significance (p = 0.014). These ex vivo findings support a myocardial locus for the contractile effect of 3‑OHB because they were obtained in a preparation isolated from systemic influences and under fixed pacing.
To test whether 3‑OHB acts through classical receptor‑mediated β‑adrenergic/cAMP pathways, the authors examined responses under pharmacologic modulation:
Coadministration of isoprenaline (a β‑adrenergic agonist) did not further augment the inotropic effect of 3‑OHB.
Blockade of β1‑adrenergic receptors with metoprolol did not attenuate the inotropic response to 3‑OHB.
Inhibition of adenylyl cyclase with 2,5‑dideoxyadenosine, limiting cAMP generation, also did not diminish the 3‑OHB–induced increase in twitch force.
These negative interaction results indicate that the inotropic action of 3‑OHB does not depend on beta‑adrenergic receptor activation or on downstream cAMP‑dependent signaling, as probed by the chosen pharmacologic tools.
Combined in vivo and ex vivo evidence indicates that 3‑OHB directly improves LV contractile performance and efficiency. The absence of enhancement by β‑adrenergic stimulation and the lack of attenuation by β1 blockade or adenylyl cyclase inhibition point away from a receptor‑mediated, cAMP‑dependent mechanism. The authors conclude that the findings are most consistent with a direct bioenergetic effect of 3‑OHB on myocardial contractility.
Accordingly, 3‑OHB is proposed as a candidate inotropic agent whose mechanistic profile differs from conventional cAMP‑dependent inotropes. The greater relative increase in twitch force observed in healthy trabeculae compared with HF tissue is reported, although both groups exhibited significant augmentation.
The work was supported by the Ministry of Health of the Czech Republic (conceptual development and institutional support for IKEM) and by Ministry of Health cooperation with the Czech Health Research Council — National Institute CarDia (NW26A‑CARDIA). The authors declared no competing interests.
This report is a preprint posted on bioRxiv and has not been certified by peer review. Details beyond those reported in the preprint (for example, full experimental protocols, sample sizes, variability measures, and extended methods or results) are available in the full preprint PDF and supplementary material referenced by the original source.