Carlos M. Villalón

Background/Objectives: Adenosine 5′-O-(β-thio)-diphosphate (ADPβS) is a stable analogue of adenosine diphosphate (ADP). In pithed rats, ADPβS produces (i) inhibition of both the cardioaccelerator sympathetic tone (via prejunctional P2Y₁₂/P2Y₁₃ receptors) and the vasodepressor sensory CGRPergic tone (via prejunctional P2Y₁ receptors) and (ii) vasodepressor/vasodilator responses via endothelial P2Y₁ receptors, and vasopressor/vasoconstrictor responses via P2Y₁/P2Y₁₂/P2Y₁₃ receptors. This study pharmacologically investigates whether P2Y₁, P2Y₁₂, and/or P2Y₁₃ receptors modulate the vasopressor sympathetic tone using ADPβS. Methods: A total of 126 male Wistar rats were pithed and classified into two groups to produce vasopressor responses by (i) spinal (T₇–T₉) electrical stimulation of the sympathetic vasopressor tone (0.03–3 Hz; n = 102) and (ii) intravenous (i.v.) boluses of noradrenaline (0.03–3 μg/kg; n = 24). Results: Continuous i.v. infusions of ADPβS (5.6 and 10 μg/kg·min) significantly inhibited the vasopressor responses to sympathetic electrical stimulation, without affecting those induced by exogenous noradrenaline. This sympathoinhibition by ADPβS (5.6 μg/kg·min) remained unchanged after vehicle (1 mL/kg, i.v.) or the antagonists PSB 0739 (P2Y_12; 300 μg/kg, i.v.) and MRS 2211 (P2Y_13; 1000 and 3000 μg/kg, i.v.). Conversely, it was abolished after MRS 2500 (P2Y_1; 300 μg/kg, i.v.) or glibenclamide (K_ATP channel blocker; 20 mg/kg, i.v.). Conclusions: ADPβS inhibits the vasopressor sympathetic tone via prejunctional P2Y₁ receptors through K_ATP channel-dependent mechanisms.

Cardiovascular modulation mediated by adenine-based purines is a very complicated and dynamic process driven by adenosine, adenosine diphosphate (ADP), and adenosine triphosphate (ATP). It is orchestrated through the interaction of these molecules with specific purinergic P1, P2Y, and P2X receptors, located throughout the central and peripheral nervous systems. This, along with the constant enzymatic interconversion of these compounds by ectonucleotidases, results in overlapping and highly variable cardiovascular responses. Activation of G protein-coupled purinergic P2Y receptors plays a fundamental role in cardiovascular homeostasis, modulating autonomic integration, neurogenic transmission, and vascular function. Within this context, ADP-sensitive P2Y receptors, particularly the P2Y1, P2Y12, and P2Y13 subtypes, participate in distinct sensory, autonomic, and functional cardiovascular mechanisms, as identified in several in vitro and in vivo experimental models. Accordingly: (i) the stable and non-hydrolysable ADP analogue, adenosine 5′-O-(β-thio)-diphosphate (ADPβS), has been widely used as a pharmacological tool to investigate P2Y receptor-mediated cardiovascular responses; and (ii) the specific role of P2Y receptors has been confirmed using selective P2Y receptor antagonists, including MRS2500 (P2Y1), PSB0739 (P2Y12) or MRS2211 (P2Y13). The cardiovascular effects of ADP are complex; depending on the experimental conditions, it can either decrease or increase heart rate, myocardial contractility, vascular tone, and/or systemic blood pressure. These effects are mediated by several P2Y receptors that act at different levels of cardiovascular control, encompassing crucial integration sites within the central nervous system, as well as peripheral autonomic and cardiovascular effector mechanisms such as preganglionic nerves, autonomic ganglia, postganglionic nerves, sensory fibres, vascular smooth muscle, vascular endothelium, and cardiac tissues. In this review, we: (i) summarize the pharmacological profile of the P2Y receptors that mediate cardiovascular responses, with special emphasis on peripheral autonomic, sensory, and vascular mechanisms, revealed in in vivo experimental models; and (ii) highlight the potential of P2Y receptors as therapeutic targets for the treatment of cardiovascular pathologies, including hypertension, coronary vasospasm, ischemic injury, and coagulation problems.

The introduction of monoclonal antibodies (mAbs) targeting the calcitonin gene-related peptide (CGRP) pathway has revolutionized migraine treatment. In this regard, galcanezumab, a humanized immunoglobulin G4 (IgG4) that binds the CGRP molecule, has rapid onset of action, monthly dosing, and favorable pharmacokinetics. This review, which draws upon electronic databases such as PubMed, Web of Science, FDA, and EMA, provides a comprehensive summary of galcanezumab for migraine prophylaxis. It covers the drug's mechanism of action, its clinical efficacy in episodic migraine, and its potential benefits in episodic cluster headache and medication-overuse headache. It also provides an overall safety profile based on clinical trials and real-world studies. Galcanezumab is a cornerstone preventive therapy with consistent efficacy, favorable tolerability, and convenient dosing. Key unmet needs include identifying biomarkers to predict responders, addressing access and cost barriers, optimizing evaluation windows for late-responders, and expanding the evidence base for special populations (pediatrics, pregnancy, and vascular comorbidities). Future studies should refine patient selection and explore complementary targets for non-responders.

Vasoinhibin is a potent antiangiogenic protein that blocks the activation of endothelial nitric oxide synthase (eNOS) in response to vascular endothelial growth factor, bradykinin, and acetylcholine (ACh). In this regard, VIAN‐c4551, a new synthetic vasoinhibin analog, has therapeutic potential for targeting angiogenesis in oncology and ophthalmology. Given that cardiovascular actions are common complications of antiangiogenic drugs and eNOS inhibitors, this multidisciplinary study investigated the cardiovascular safety of VIAN‐c4551. Administered acutely, VIAN‐c4551 inhibited ACh‐induced eNOS phosphorylation/activation in cultured endothelial cells and in lung tissue treated in vivo, as well as the ACh‐induced relaxation of rat aortic segments. However, daily intravenous (i.v.) injections of VIAN‐c4551 (1 or 3 mg kg ⁻¹ ) for 5 days failed to significantly modify the vasodepressor responses to ACh and the baseline values of blood pressure in anesthetized rats (intact, vagotomized, or pithed). Furthermore, daily i.v. injections of 1 mg kg ⁻¹ VIAN‐c4551 (for 5 days) did not alter: (i) blood pressure or heart rate values in awake rats; (ii) cardiac autonomic and histological outcomes in anesthetized animals; or (iii) inflammatory (tumor necrosis factor‐α [TNF‐α] and interleukin‐6 [IL‐6]) and apoptotic (caspase‐3) markers. Although VIAN‐c4551 inhibited ACh‐induced eNOS activation in vitro and in vivo and vasorelaxation in ex vivo assays, in vivo experiments consistently showed no significant cardiovascular effects produced by this synthetic peptide. Thus, VIAN‐c4551 appears to be cardiovascularly safe for targeting angiogenesis‐related diseases.

Background and Purpose: Sex‐dependent vascular effects of transient receptor potential (TRP) channels and sex dimorphism in migraine are not yet fully characterized. We investigated the differential vasoactive effects of TRP ankyrin 1 (TRPA1), TRP melastatin 3 (TRPM3) and TRP vanilloid 1 (TRPV1) channels, their pharmacological mechanism(s), and localization and expression in human isolated blood vessels.Experimental Approach: Agonist responses to cinnamaldehyde (TRPA1), pregnenolone sulfate (PregS, TRPM3) or capsaicin (TRPV1) were analysed using wire myography in segments of human coronary (HCAs) and middle meningeal (HMMAs) arteries from men and women. The mechanisms involved in these responses were investigated using the antagonists/blockers/inhibitors: HC‐030031 (TRPA1), isosakuranetin (TRPM3), capsazepine (TRPV1), olcegepant (calcitonin gene‐related peptide [CGRP] receptor), L‐NAME (nitric oxide synthase [NOS]), indomethacin (cyclooxygenase [COX]), TRAM‐34 + apamin (K⁺ channels) or MK‐801 (N‐methyl‐d‐aspartate [NMDA] receptor). Fluorescence microscopy, quantitative polymerase chain reaction (qPCR), and western blotting were performed to investigate their location and expression, respectively.Key Results: In HCAs and HMMAs, (i) capsaicin‐induced relaxation remained unchanged after the above‐mentioned antagonists/blockers/inhibitors and (ii) cinnamaldehyde‐induced relaxation was blocked by olcegepant. PregS‐induced maximal relaxation was significantly enhanced in isolated arteries from females compared with males and was inhibited after isosakuranetin, MK‐801 or L‐NAME. TRPM3 mRNA and protein expression, along with NMDA protein levels, were higher in arteries from females than males.Conclusion and Implications: Modulation of vascular tone in HCAs and HMMAs by activation of TRPM3 is sex‐dependent, likely involving NMDA receptors. This represents a new therapeutic direction, targeting sex dimorphism in migraine and its related cardiovascular events.

Migraine is a complex disorder, and its etiology is not yet fully understood. In the last 40 years, calcitonin gene-related peptide (CGRP) has been central to the understanding of migraine pathophysiology, leading to the development of new molecules targeting the CGRPergic system. These new molecules, such as gepants and monoclonal antibodies, are effective, well-tolerated, and safe, and are approved for clinical use. By searching multiple electronic scientific databases, this narrative review examined: (i) the role of CGRP in migraine; and (ii) the current knowledge on the effects of CGRPergic antimigraine pharmacotherapies, including a brief analysis of their pharmacodynamic and pharmacokinetic characteristics. Current anti-CGRPergic medications, although effective, have limitations, such as side effects and lack of antimigraine efficacy in some patients. The existence of patients with medication-resistant migraine may be due to the: (i) complex migraine pathophysiology, in which several systems appear to be deregulated before, during, and after a migraine attack; and (ii) pharmacodynamic and pharmacokinetic properties of antimigraine medications. As envisioned here, although seminal studies support the notion that CGRP plays a key role in migraine headache, the dysfunction of CGRPergic transmission does not seem to be relevant in all cases.

Glyphosate, a widely used herbicide, is linked to a plethora of deleterious effects in both clinical and preclinical studies. Nevertheless, the effects of its main metabolite, aminomethylphosphonic acid (AMPA), whose half-life in soil is even longer than that of glyphosate, have been little explored. On this basis, as a first approach, in this work, we report that intraperitoneal (i.p.) administration of AMPA or glyphosate (at 10, 56, and 100 mg/kg) decreased, to a similar extent, plasma cholinesterase (ChE) activity in acutely exposed rats. Moreover, we designed an experimental protocol to analyze and compare the effects of AMPA and glyphosate on human plasma ChE activity; this protocol consisted of adding these compounds to human plasma to subsequently test the effects of this plasma on the contraction to acetylcholine (ACh) in the frog rectus abdominis muscle (an indirect estimate of ChE activity). Accordingly, this muscular contraction to ACh was evaluated before and after pre-incubation of ACh with (i) plasma alone, (ii) plasma with AMPA, and (iii) plasma with glyphosate. Our results indicate that AMPA, like glyphosate, decreased ChE activity in the plasma of rats (when given i.p.) and humans (when added in vitro), suggesting that both xenobiotics may exert similar toxicological effects.

Glyphosate, a widely used herbicide, is linked to a plethora of deleterious effects in both clinical and preclinical studies. Nevertheless, the effects of its main metabolite, aminomethylphosphonic acid (AMPA), whose half-life in soil is even longer than that of glyphosate, have been little explored. On this basis, as a first approach, in this work we report that intraperitoneal (i.p.) administration of AMPA or glyphosate (at 10, 56, and 100 mg/kg) decreased, to a similar extent, plasma cholinesterase (ChE) activity in acutely exposed rats. Moreover, we designed an experimental protocol to analyze and compare the effects of AMPA and glyphosate on human plasma ChE activity; this protocol consisted of adding these compounds to human plasma to subsequently test the effects of this plasma on the contraction to acetylcholine (ACh) in the frog rectus abdominis muscle (an indirect estimate of ChE activity). Accordingly, this muscular contraction to ACh was evaluated before and after pre‑incubation of ACh with: (i) plasma alone; (ii) plasma with AMPA; and (iii) plasma with glyphosate. Our results indicate that AMPA, like glyphosate, decreased ChE activity in the plasma of rats (when given i.p.) and humans (when added in vitro ), suggesting that both xenobiotics may exert similar toxicological effects.

Transient receptor potential (TRP) channels are pivotal in modulating vascular functions. In fact, topical application of cinnamaldehyde or capsaicin (TRPA1 and TRPV1 channel agonists, respectively) induces “local” changes in blood flow by releasing vasodilator neuropeptides. We investigated TRP channels’ contributions and the pharmacological mechanisms driving vasodilation in human isolated dermal arteries. Ex vivo studies assessed the vascular function of artery segments and analyzed the effects of different compounds. Concentration–response curves to cinnamaldehyde, pregnenolone sulfate (PregS, TRPM3 agonist), and capsaicin were constructed to evaluate the effect of the antagonists HC030031 (TRPA1); isosakuranetin (TRPM3); and capsazepine (TRPV1). Additionally, the antagonists/inhibitors olcegepant (CGRP receptor); L-NAME (nitric oxide synthase); indomethacin (cyclooxygenase); TRAM-34 plus apamin (K⁺ channels); and MK-801 (NMDA receptors, only for PregS) were used. Moreover, CGRP release was assessed in the organ bath fluid post-agonist-exposure. In dermal arteries, cinnamaldehyde- and capsaicin-induced relaxation remained unchanged after the aforementioned antagonists, while PregS-induced relaxation was significantly inhibited by isosakuranetin, L-NAME and MK-801. Furthermore, there was a significant increase in CGRP levels post-agonist-exposure. In our experimental model, TRPA1 and TRPV1 channels seem not to be involved in cinnamaldehyde- or capsaicin-induced relaxation, respectively, whereas TRPM3 channels contribute to PregS-induced relaxation, possibly via CGRP-independent mechanisms.

Purine nucleosides (adenosine) and nucleotides such as adenosine mono/di/triphosphate (AMP/ADP/ATP) may produce complex cardiovascular responses. For example, adenosine-5′-(β-thio)-diphosphate (ADPβS; a stable synthetic analogue of ADP) can induce vasodilatation/vasodepressor responses by endothelium-dependent and independent mechanisms involving purinergic P2Y receptors; however, the specific subtypes participating in these responses remain unknown. Therefore, this study investigated the receptor subtypes mediating the blood pressure changes induced by intravenous bolus of ADPβS in male Wistar rats in the absence and presence of central mechanisms with the antagonists MRS2500 (P2Y₁), PSB0739 (P2Y₁₂), and MRS2211 (P2Y₁₃). For this purpose, 120 rats were divided into 60 anaesthetised rats and 60 pithed rats, and further subdivided into four groups (n = 30 each), namely: (a) anaesthetised rats, (b) anaesthetised rats with bilateral vagotomy, (c) pithed rats, and (d) pithed rats continuously infused (intravenously) with methoxamine (an α₁-adrenergic agonist that restores systemic vascular tone). We observed, in all four groups, that the immediate decreases in diastolic blood pressure produced by ADPβS were exclusively mediated by peripheral activation of P2Y₁ receptors. Nevertheless, the subsequent increases in systolic blood pressure elicited by ADPβS in pithed rats infused with methoxamine probably involved peripheral activation of P2Y₁, P2Y₁₂, and P2Y₁₃ receptors.