Spontaneous calcium waves in cardiac myocytes are caused by diastolic sarcoplasmic reticulum release (SR Ca²⁺ leak) through ryanodine receptors. Beta-adrenergic (β-AR) tone is known to increase this leak through the activation of Ca-calmodulin-dependent protein kinase (CaMKII) and the subsequent phosphorylation of the ryanodine receptor. When β-AR drive is chronic, as observed in heart failure, this CaMKII-dependent effect is exaggerated and becomes potentially arrhythmogenic. Recent evidence has indicated that CaMKII activation can be regulated by cellular oxidizing agents, such as reactive oxygen species. Here, we investigate how the cellular second messenger, nitric oxide, mediates CaMKII activity downstream of the adrenergic signaling cascade and promotes the generation of arrhythmogenic spontaneous Ca²⁺ waves in intact cardiomyocytes. Both SCaWs and SR Ca²⁺ leak were measured in intact rabbit and mouse ventricular myocytes loaded with the Ca-dependent fluorescent dye, fluo-4. CaMKII activity in vitro and immunoblotting for phosphorylated residues on CaMKII, nitric oxide synthase, and Akt were measured to confirm activity of these enzymes as part of the adrenergic cascade. We demonstrate that stimulation of the β-AR pathway by isoproterenol increased the CaMKII-dependent SR Ca²⁺ leak. This increased leak was prevented by inhibition of nitric oxide synthase 1 but not nitric oxide synthase 3. In ventricular myocytes isolated from wild-type mice, isoproterenol stimulation also increased the CaMKII-dependent leak. Critically, in myocytes isolated from nitric oxide synthase 1 knock-out mice this effect is ablated. We show that isoproterenol stimulation leads to an increase in nitric oxide production, and nitric oxide alone is sufficient to activate CaMKII and increase SR Ca²⁺ leak. Mechanistically, our data links Akt to nitric oxide synthase 1 activation downstream of β-AR stimulation. Collectively, this evidence supports the hypothesis that CaMKII is regulated by nitric oxide as part of the adrenergic cascade leading to arrhythmogenesis.

Secretin is a 27-amino acid brain-gut peptide from duodenal S-cells. We tested the effects of systemic administration of secretin to simulate its postprandial release on neuroendocrine neurons of the supraoptic nucleus (SON) in urethane-anesthetized female rats. Secretin dose-dependently increased the firing rate of oxytocin neurons, more potently than cholecystokinin, and dose-dependently increased plasma oxytocin concentration. The effect of secretin on SON vasopressin neurons was also predominantly excitatory, in contrast to the inhibitory actions of cholecystokinin. To explore the involvement of noradrenergic inputs in secretin-induced excitation, benoxathian, an α1-adrenoceptor antagonist, was infused intracerebroventricularly. Benoxathian intracerebroventricular infusion blocked the excitation by secretin of both oxytocin and vasopressin neurons. To test the role of local noradrenaline release in the SON, benoxathian was microdialyzed onto the SON. The basal firing rate of oxytocin neurons was slightly reduced and the secretin-induced excitation was attenuated during benoxathian microdialysis. Hence, noradrenergic pathways mediate the excitation by systemic secretin of oxytocin neurons via α1-adrenoceptors in the SON. As both systemic secretin and oxytocin are involved in regulating gastrointestinal functions and natriuresis, systemically released secretin might act partly through oxytocin.

Background: Mitochondrial dysfunction contributes to the cardiac remodeling triggered by type 2 diabetes (T2D). Mitochondrial Ca ²⁺ concentration ([Ca ²⁺ ] _m ) modulates the oxidative state and cytosolic Ca ²⁺ regulation. Thus, we investigated how T2D affects mitochondrial Ca ²⁺ fluxes, the downstream consequences on myocyte function, and the effects of normalizing mitochondrial Ca ²⁺ transport. Methods and Results: We compared myocytes/hearts from transgenic rats with late‐onset T2D (rats that develop late‐onset T2D due to heterozygous expression of human amylin in the pancreatic β‐cells [HIP] model) and their nondiabetic wild‐type (WT) littermates. [Ca ²⁺ ] _m was significantly lower in myocytes from diabetic HIP rats compared with WT cells. Ca ²⁺ extrusion through the mitochondrial Na ⁺ /Ca ²⁺ exchanger (mitoNCX) was elevated in HIP versus WT myocytes, particularly at moderate and high [Ca ²⁺ ] _m , while mitochondrial Ca ²⁺ uptake was diminished. Mitochondrial Na ⁺ concentration was comparable in WT and HIP rat myocytes and remained remarkably stable while manipulating mitoNCX activity. Lower [Ca ²⁺ ] _m was associated with oxidative stress, increased sarcoplasmic reticulum Ca ²⁺ leak in the form of Ca ²⁺ sparks, and mitochondrial dysfunction in T2D hearts. MitoNCX inhibition with CGP‐37157 reduced oxidative stress, Ca ²⁺ spark frequency, and stress‐induced arrhythmias in HIP rat hearts while having no significant effect in WT rats. In contrast, activation of the mitochondrial Ca ²⁺ uniporter with SB‐202190 enhanced spontaneous sarcoplasmic reticulum Ca ²⁺ release and had no significant effect on arrhythmias in both WT and HIP rat hearts. Conclusions: [Ca ²⁺ ] _m is reduced in myocytes from rats with T2D due to a combination of exacerbated mitochondrial Ca ²⁺ extrusion through mitoNCX and impaired mitochondrial Ca ²⁺ uptake. Partial mitoNCX inhibition limits sarcoplasmic reticulum Ca ²⁺ leak and arrhythmias in T2D hearts, whereas mitochondrial Ca ²⁺ uniporter activation does not.

Kisspeptin is a neuropeptide that governs the reproductive axis upstream to GnRH. We wanted to study whether kisspeptin modulates plasma LH and FSH levels and ovarian follicular dynamics in buffaloes and whether kisspeptin can be used for fixed time artificial insemination (FTAI). We carried out these studies in comparison with buserelin, a potent GnRH agonist. Kisspeptin dose-dependently increased plasma LH levels. However, the kisspeptin-induced increase in LH was short-lived as the peak reached in 15–30 min returned to basal values by 1–2 h. The kisspeptin-induced increase in LH level was less compared to buserelin-induced increase in LH level which sustained over time. Kisspeptin did not enhance FSH release while buserelin resulted in a gradual increase over time. LH response to repeated injections of kisspeptin was greater than that induced by buserelin. While buserelin induced an increase in the number of follicles, kisspeptin induced an increase in the growth rate of the follicle. In adult cycling animals, while both the drugs increased plasma LH levels, the increase was greater in buserelin group compared to kisspeptin group. In contrast to the findings in pre-pubertal animals, kisspeptin induced an increase in both the number as well as the size of follicles compared to buserelin. Our studies on oestrus synchronization, using either kisspeptin-PGF_2α-kisspeptin protocol or buserelin-PGF_2α-buserelin Ovsynch protocol on day 0, 7, and 9, respectively, revealed that kisspeptin increased the number of follicles at wave emergence and the diameter of dominant follicle after 2nd dose of drug, the oestrus response rate and duration of oestrus, compared to buserelin. However, conception rate was not significantly different among the groups. From our studies, it appears that Kp and Buserelin differentially modulate follicular dynamics depending on the reproductive age of the animals.However, studies in a larger herd are required to confirm whether kisspeptin can be used for oestrous synchronization in buffaloes.