British Journal of Pharmacology最新文献

Background and purpose: Fentanyl analogues have been implicated in many cases of intoxication and death with overdose worldwide. The aim of this study is to investigate the pharmaco-toxicology of two fentanyl analogues: butyrylfentanyl (BUF) and 4-fluorobutyrylfentanyl (4F-BUF).

Experimental approach: In vitro, we measured agonist opioid receptor efficacy, potency, and selectivity and ability to promote interaction of the μ receptor with G protein and β-arrestin 2. In vivo, we evaluated thermal antinociception, stimulated motor activity and cardiorespiratory changes in female and male CD-1 mice injected with BUF or 4F-BUF (0.1-6 mg·kg^-1). Opioid receptor specificity was investigated using naloxone (6 mg·kg^-1). We investigated the possible role of stress in increasing cardiorespiratory toxicity using the corticotropin-releasing factor 1 (CRF₁) antagonist antalarmin (10 mg·kg^-1).

Key results: Agonists displayed the following rank of potency at μ receptors: fentanyl > 4F-BUF > BUF. Fentanyl and BUF behaved as partial agonists for the β-arrestin 2 pathway, whereas 4F-BUF did not promote β-arrestin 2 recruitment. In vivo, we revealed sex differences in motor and cardiorespiratory impairments but not antinociception induced by BUF and 4F-BUF. Antalarmin alone was effective in blocking respiratory impairment induced by BUF in both sexes but not 4F-BUF. The combination of naloxone and antalarmin significantly enhanced naloxone reversal of the cardiorespiratory impairments induced by BUF and 4F-BUF in mice.

Conclusion and implications: In this study, we have uncovered a novel mechanism by which synthetic opioids induce respiratory depression, shedding new light on the role of CRF₁ receptors in cardiorespiratory impairments by μ agonists.

Background and purpose: Transient receptor potential vanilloid 2 (TRPV2) is a Ca²⁺-permeable non-selective cation channel. Despite the significant roles of TRPV2 in immunological response, cancer progression and cardiac development, pharmacological probes of TRPV2 remain to be identified. We aimed to discover TRPV2 inhibitors and to elucidate their molecular mechanism of action.

Experimental approach: Fluorescence-based Ca²⁺ assay in HEK-293 cells expressing murine TRPV2 was used to identify plumbagin as a novel TRPV2 inhibitor. Patch-clamp, in silico docking and site-directed mutagenesis were applied to investigate the molecular mechanisms critical for plumbagin interaction. ELISA and qPCR were used to assess nitric oxide release and mRNA levels of inflammatory mediators, respectively. si-RNA interference was used to knock down TRPV2 expression, which was validated by western blotting. Neurological and histological analyses were used to examine brain injury of mice following middle cerebral artery occlusion/reperfusion (MCAO/R).

Key results: Plumbagin is a potent TRPV2 negative allosteric modulator with an IC₅₀ value of 0.85 μM, exhibiting >14-fold selectivity over TRPV1, TRPV3 and TRPV4. Plumbagin suppresses TRPV2 activity by decreasing the channel open probability without affecting the unitary conductance. Moreover, plumbagin binds to an extracellular pocket formed by the pore helix and flexible loop between transmembrane helices S5 and S6 of TRPV2. Plumbagin effectively suppresses LPS-induced inflammation of BV-2 microglia and ameliorates brain injury of MCAO/R mice.

Conclusion and implications: Plumbagin is a novel pharmacological probe to study TRPV2 pathophysiology. TRPV2 is a novel molecular target for the treatment of neuroinflammation and ischemic stroke.

Background and purpose: GABAergic neurons in mouse ventral tegmental area (VTA) exhibit elevated activity during withdrawal following chronic ethanol exposure. While increased glutamatergic input and decreased GABA_A receptor sensitivity have been implicated, the impact of inhibitory signaling in VTA GABA neurons has not been fully addressed.

Experimental approach: We used electrophysiological and ultrastructural approaches to assess the impact of chronic intermittent ethanol vapour exposure in mice on GABAergic transmission in VTA GABA neurons during withdrawal. We used CRISPR/Cas9 ablation to mimic a somatodendritic adaptation involving the GABA_B receptor (GABA_BR) in ethanol-naïve mice to investigate its impact on anxiety-related behaviour.

Key results: The frequency of spontaneous inhibitory postsynaptic currents was reduced in VTA GABA neurons following chronic ethanol treatment and this was reversed by GABA_BR inhibition, suggesting chronic ethanol strengthens the GABA_BR-dependent suppression of GABAergic input to VTA GABA neurons. Similarly, paired-pulse depression of GABA_A receptor-dependent responses evoked by optogenetic stimulation of nucleus accumbens inputs from ethanol-treated mice was reversed by GABA_BR inhibition. Somatodendritic currents evoked in VTA GABA neurons by GABA_BR activation were reduced following ethanol exposure, attributable to the suppression of GIRK (K_ir3) channel activity. Mimicking this adaptation enhanced anxiety-related behaviour in ethanol-naïve mice.

Conclusions and implications: Chronic ethanol weakens the GABAergic regulation of VTA GABA neurons in mice via pre- and postsynaptic mechanisms, likely contributing to their elevated activity during withdrawal and expression of anxiety-related behaviour. As anxiety can promote relapse during abstinence, interventions targeting VTA GABA neuron excitability could represent new therapeutic strategies for treatment of alcohol use disorder.

This brief review highlights some of the structure-activity relationships of classic serotonergic psychedelics. In particular, we discuss structural features of three chemotypes: phenethylamines, ergolines and certain tryptamines, which possess psychedelic activity in humans. Where they are known, we point out the underlying molecular mechanisms utilized by each of the three chemotypes of psychedelic molecules. With a focus on the 5-HT_2A receptor subtype, a G-protein coupled receptor known to be the primary target of psychedelics, we refer to several X-ray and cryoEM structures, with a variety of ligands bound, to illustrate the underlying atomistic basis for some of the known pharmacological observations of psychedelic drug actions.

Background and purpose: Diabetic nephropathy (DN) is a leading cause of chronic kidney disease (CKD), which is characterized by mesangial matrix expansion that involves dysfunctional mesangial cells (MCs). However, the underlying mechanisms remain unclear. This study aims to delineate the spatiotemporal contribution of adrenergic signalling in diabetic kidney fibrosis to reveal potential therapeutic targets.

Experimental approach: A model of diabetic nephropathy was induced by in db/db mice. Gene expression in kidneys was profiled by RNA-seq analyses, western blot and immunostaining. Subcellular-localized fluorescence resonance energy transfer (FRET) biosensors determined adrenergic signalling microdomains in MCs. Effects of oral rolipram, a phosphodiesterase 4 (PDE4) inhibitor, on the model were measured.

Key results: Our model exhibited impaired kidney function with elevated expression of adrenergic and fibrotic genes, including Adrb1, PDEs, Acta2 and Tgfβ. RNA-seq analysis revealed that MCs with dysregulated YAP pathway were crucial to the extracellular matrix secretion in kidneys from diabetic nephropathy patients. In cultured MCs, TGF-β promoted profibrotic gene transcription, which was regulated by nuclear-localized β-adrenoceptor signalling. Mechanistically, TGF-β treatment diminished nuclear-specific cAMP signalling in MCs and reduced PKA-dependent phosphorylation of YAP, leading to its activation. In parallel, db/db mouse kidneys showed increased expressions of PDE4B and PDE4D. Treatment with oral rolipram alleviated kidney fibrosis in db/db mice.

Conclusion and implications: Diabetic nephropathy impaired nuclear-localized β₁-adrenoceptor-cAMP signalling microdomain through upregulating PDE4 expression, promoting fibrosis in MCs via PKA dephosphorylation-dependent YAP activation. Our results suggest PDE4 inhibition as a promising strategy for alleviating kidney fibrosis in diabetic nephropathy.

Background and purpose: The pharmacology of flavonoids on β-cell function is largely undefined especially in the context of defective secretion of insulin. We sought to identify flavonoids that increased the insulin-secreting function of β-cells and to explore the underlying mechanisms.

Experimental approach: INS-1 β-cells in culture and islets of Langerhans isolated from control and diabetic male rats were used for insulin secretion experiments. Pharmacological and electrophysiological approaches were used for mechanistic studies.

Key results: Among a set of flavonoids, exposure of INS-1 β-cells to resokaempferol (ResoK) enhanced glucose-stimulated insulin secretion and therefore we further characterised its activity and its pharmacological mechanism. ResoK glucose-dependently enhanced insulin secretion in INS-1 β-cells and pancreatic islets isolated from rats. Mechanistically, whole cell patch clamp recordings in INS-1 cells showed that ResoK rapidly and dose-dependently enhanced the L-type Ca²⁺ current whereas it was inactive towards T-type Ca²⁺ current. Accordingly, pharmacological inhibition of L-type Ca²⁺ current but not T-type Ca²⁺ current blocked the effects of ResoK on glucose-stimulated insulin secretion. ResoK was still active on dysfunctional β-cells as it ameliorated glucose-stimulated insulin secretion in glucotoxicity-induced dysfunctional INS-1 cells and in pancreatic islets isolated from diabetic rats.

Conclusion and implications: ResoK is a glucose-dependent activator of insulin secretion. Our results indicated that the effects of ResoK on insulin secretion involved its capacity to stimulate L-type Ca²⁺ currents in cultured β-cells. As ResoK was also effective on dysfunctional β-cells, our work provides a new approach to stimulating insulin secretion, using compounds based on the structure of ResoK.

The orexin (also known as hypocretin) system, consisting of neuropeptides orexin-A and orexin-B, was discovered over 25 years ago and was immediately identified as a central regulator of sleep and wakefulness. These peptides interact with two G-protein coupled receptors, orexin 1 (OX₁) and orexin 2 (OX₂) receptors which are capable of coupling to all heterotrimeric G-protein subfamilies, but primarily transduce increases in calcium signalling. Orexin neurons are regulated by a variety of transmitter systems and environmental stimuli that signal reward availability, including food and drug related cues. Orexin neurons are also activated by anticipation, stress, cues predicting motivationally relevant information, including those predicting drugs of abuse, and engage neuromodulatory systems, including dopamine neurons of the ventral tegmental area (VTA) to respond to these signals. As such, orexin neurons have been characterized as motivational activators that coordinate a range of functions, including feeding and arousal, that allow the individual to respond to motivationally relevant information, critical for survival. This review focuses on the role of orexins in appetitive motivation and highlights a role for these neuropeptides in pathologies characterized by inappropriately high levels of motivated arousal (overeating, anxiety and substance use disorders) versus those in which motivation is impaired (depression).