Utilising fluorescent reporters to probe the mode of action of norbornen-7-one CO releasing molecules.

The synthesis of the fluorescent organic carbon monoxide releasing molecules oCOm-57, oCOm-58, and oCOm-66 are reported. These oCOms are water soluble and exhibit a "turn-on" fluorescent behaviour when CO is released under physiological conditions. oCOm-66 also contains an additional nitro-naphthalimide moiety that functions as a fluorescent reporter. Delivery of CO released from these oCOms to the mitochondria of AC-16 cardiomyocytes was confirmed using confocal microscopy in conjuction with MitoTracker Red. While the neutral, PEGylated oCOm-57 was found to remain in the extracellular environment releasing CO to diffuse into the cellular compartments, the positively charged oCOm-58 and -66 are targeted to the mitochondria where they release CO. Notably, the use of the fluorescent oCOms in live cellular imaging, allows the intracellular CO delivery and oCOm localisation to be characterised. This cellular confocal study also shows that, subtoxic concentrations of CO released from these molecules preserved mitochondrial energetics as indicated by the membrane potential dependent MitoTracker Red.

A Clinically Relevant Functional Model of Type-2 Cardio-Renal Syndrome with Paraventricular Changes consequent to Chronic Ischaemic Heart Failure.

Cardiorenal syndrome, de novo renal pathology arising secondary to cardiac insufficiency, is clinically recognised but poorly characterised. This study establishes and characterises a valid model representative of Type 2 cardiorenal syndrome. Extensive permanent left ventricular infarction, induced by ligation of the left anterior descending coronary artery in Lewis rats, was confirmed by plasma cardiac troponin I, histology and cardiac haemodynamics. Renal function and morphology was assessed 90-days post-ligation when heart failure had developed. The involvement of the paraventricular nucleus was investigated using markers of inflammation, apoptosis, reactive oxygen species and of angiotensin II involvement. An extensive left ventricular infarct was confirmed following coronary artery ligation, resulting in increased left ventricular weight and compromised left ventricular diastolic function and developed pressure. Glomerular filtration was significantly decreased, fractional excretion of sodium and caspase activities were increased and basement membrane thickening, indicating glomerulosclerosis, was evident. Interestingly, angiotensin II receptor I expression and reactive oxygen species levels in the hypothalamic paraventricular nucleus remained significantly increased at 90-days post-coronary artery ligation, suggesting that these hypothalamic changes may represent a novel, valuable pharmacological target. This model provides conclusive morphological, biochemical and functional evidence of renal injury consequent to heart failure, truly representative of Type-2 cardiorenal syndrome.