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Journal of the... 2020Amiloride is a potassium retaining diuretic and natriuretic which acts by reversibly blocking luminal epithelial sodium channels (ENaCs) in the late distal tubule and... (Review)
Review
Amiloride is a potassium retaining diuretic and natriuretic which acts by reversibly blocking luminal epithelial sodium channels (ENaCs) in the late distal tubule and collecting duct. Amiloride is indicated in oedematous states, and for potassium conservation adjunctive to thiazide or loop diuretics for hypertension, congestive heart failure and hepatic cirrhosis with ascites. Historical studies on its use in hypertension were poorly controlled and there is insufficient data on dose-response. It is clearly highly effective in combination with thiazide diuretics where it counteracts the adverse metabolic effects of the thiazides and its use in the Medical Research Council Trial of Older Hypertensive Patients, demonstrated convincing outcome benefits on stroke and coronary events. Recently it has been shown to be as effective as spironolactone in resistant hypertension but there is a real need to establish its potential role in the much larger number of patients with mild to moderate hypertension in whom there is a paucity of information with amiloride particularly across an extended dose range.
Topics: Amiloride; Animals; Clinical Trials as Topic; Diuretics; Humans; Hypertension
PubMed: 33234024
DOI: 10.1177/1470320320975893 -
Nephrology, Dialysis, Transplantation :... Jan 2022
Topics: Amiloride; Calcium; Humans
PubMed: 34264323
DOI: 10.1093/ndt/gfab221 -
Medicinal Research Reviews Mar 2020The function of G protein-coupled receptors (GPCRs) can be modulated by compounds that bind to other sites than the endogenous orthosteric binding site, so-called... (Review)
Review
The function of G protein-coupled receptors (GPCRs) can be modulated by compounds that bind to other sites than the endogenous orthosteric binding site, so-called allosteric sites. Structure elucidation of a number of GPCRs has revealed the presence of a sodium ion bound in a conserved allosteric site. The small molecule amiloride and analogs thereof have been proposed to bind in this same sodium ion site. Hence, this review seeks to summarize and reflect on the current knowledge of allosteric effects by amiloride and its analogs on GPCRs. Amiloride is known to modulate adenosine, adrenergic, dopamine, chemokine, muscarinic, serotonin, gonadotropin-releasing hormone, GABA , and taste receptors. Amiloride analogs with lipophilic substituents tend to be more potent modulators than amiloride itself. Adenosine, α-adrenergic and dopamine receptors are most strongly modulated by amiloride analogs. In addition, for a few GPCRs, more than one binding site for amiloride has been postulated. Interestingly, the nature of the allosteric effect of amiloride and derivatives varies considerably between GPCRs, with both negative and positive allosteric modulation occurring. Since the sodium ion binding site is strongly conserved among class A GPCRs it is to be expected that amiloride also binds to class A GPCRs not evaluated yet. Investigating this typical amiloride-GPCR interaction further may yield general insight in the allosteric mechanisms of GPCR ligand binding and function, and possibly provide new opportunities for drug discovery.
Topics: Allosteric Regulation; Amiloride; Animals; Drug Discovery; Humans; Receptors, G-Protein-Coupled
PubMed: 31495942
DOI: 10.1002/med.21633 -
International Journal of Molecular... Mar 1999Amiloride and its derivatives are important tools for studying NHE-1, the ubiquitous isoform of the sodium/hydrogen exchanger protein family. Three residues in putative... (Review)
Review
Amiloride and its derivatives are important tools for studying NHE-1, the ubiquitous isoform of the sodium/hydrogen exchanger protein family. Three residues in putative transmembrane domains IV and IX have been implicated in amiloride binding and several models of the proposed amiloride-binding site have been reported. Though it has been shown that sodium ions and amiloride molecules interact at unique regions of the NHE-1 protein, physiological experiments reveal a competitive relationship between the two under some circumstances. The two binding sites are thus on closely related but distinct regions on the protein.
Topics: Amiloride; Amino Acid Sequence; Animals; Binding Sites; Humans; Membrane Proteins; Molecular Sequence Data; Protein Binding; Protein Isoforms; Protein Structure, Tertiary; Sodium; Sodium-Hydrogen Exchangers
PubMed: 10028059
DOI: 10.3892/ijmm.3.3.315 -
PloS One 2020Anxiety disorders (AD) are the most common mental conditions affecting an estimated 40 million adults in the United States. Amiloride, a diuretic agent, has shown...
Anxiety disorders (AD) are the most common mental conditions affecting an estimated 40 million adults in the United States. Amiloride, a diuretic agent, has shown efficacy in reducing anxious responses in preclinical models by inhibiting the acid-sensing ion channels (ASIC). By delivering amiloride via nasal route, rapid onset of action can be achieved due to direct "nose-to-brain" access. Therefore, this study reports the formulation, physical, chemical, and microbiological stability of an extemporaneously prepared amiloride 2 mg/mL nasal spray. The amiloride nasal spray was prepared by adding 100 mg of amiloride hydrochloride to 50 mL of sterile water for injection in a sterile reagent bottle. A stability-indicating high-performance liquid chromatography (HPLC) method was developed and validated. Forced-degradation studies were performed to confirm the ability of the HPLC method to identify the degradation products from amiloride distinctively. The physical stability of the amiloride nasal spray was assessed by pH, clarity, and viscosity assessments. For chemical stability studies, samples of nasal sprays stored at room temperature were collected at time-points 0, 3 hr., 24 hr., and 7 days and were assayed in triplicate using the stability-indicating HPLC method. Microbiological stability of the nasal spray solution was evaluated for up to 7 days based on the sterility test outlined in United States Pharmacopoeia (USP) chapter 71. The stability-indicating HPLC method identified the degradation products of amiloride without interference from amiloride. All tested solutions retained over 90% of the initial amiloride concentration for the 7-day study period. There were no changes in color, pH, and viscosity in any sample. The nasal spray solutions were sterile for up to 7 days in all samples tested. An extemporaneously prepared nasal spray solution of amiloride hydrochloride (2 mg/mL) was physically, chemically, and microbiologically stable for 7 days when stored at room temperature.
Topics: Amiloride; Drug Compounding; Drug Stability; Drug Storage; Nasal Sprays
PubMed: 32649677
DOI: 10.1371/journal.pone.0232435 -
European Journal of Pharmacology Jan 2024The use of morphine in clinical medicine is severely constrained by tolerance. Therefore, it is essential to examine pharmacological therapies that suppress the...
BACKGROUND
The use of morphine in clinical medicine is severely constrained by tolerance. Therefore, it is essential to examine pharmacological therapies that suppress the development of morphine tolerance. Amiloride suppressed the expression of inflammatory cytokines by inhibiting microglial activation. Microglia play a crucial role in the establishment of morphine tolerance. Thus, we anticipated that amiloride might suppress the development of morphine tolerance. During this investigation, we assessed the impact of amiloride on mouse morphine tolerance.
METHODS
Mice received morphine (10 mg/kg, s.c.) twice daily with intrathecally injected amiloride (0.3 μg/5 μl, 1 μg/5 μl, and 3 μg/5 μl) for nine continuous days. To assess morphine tolerance, mice underwent the tail-flick and hot plate tests. BV-2 cells were used to investigate the mechanism of amiloride. By using Western blotting, real-time PCR, and immunofluorescence labeling methods, the levels of acid-sensing ion channels (ASICs), nuclear factor kappa B (NF-kB) p65, p38 mitogen-activated protein kinase (MAPK) proteins, and neuroinflammation-related cytokines were determined.
RESULTS
The levels of ASIC3 in the spinal cord were considerably increased after long-term morphine administration. Amiloride was found to delay the development of tolerance to chronic morphine assessed via tail-flick and hot plate tests. Amiloride reduced microglial activation and downregulated the cytokines IL-1β and TNF-a by inhibiting ASIC3 in response to morphine. Furthermore, amiloride reduced p38 MAPK phosphorylation and inhibited NF-κB expression.
CONCLUSIONS
Amiloride effectively reduces chronic morphine tolerance by suppressing microglial activation caused by morphine by inhibiting ASIC3.
Topics: Mice; Animals; Morphine; Analgesics, Opioid; Amiloride; Neuroinflammatory Diseases; NF-kappa B; Microglia; Cytokines; Spinal Cord
PubMed: 37918499
DOI: 10.1016/j.ejphar.2023.176173 -
CNS Neuroscience & Therapeutics Jun 2016ASIC1a, the predominant acid-sensing ion channels (ASICs), is implicated in neurological disorders including stroke, traumatic spinal cord injury, and ALS. Potent ASIC1a...
BACKGROUND
ASIC1a, the predominant acid-sensing ion channels (ASICs), is implicated in neurological disorders including stroke, traumatic spinal cord injury, and ALS. Potent ASIC1a inhibitors should have promising therapeutic potential for ASIC1a-related diseases.
AIMS
We examined the inhibitory effects of a number of amiloride analogs on ASIC1a currents, aimed at understanding the structure-activity relationship and identifying potent ASIC1a inhibitors for stroke intervention.
METHODS
Whole-cell patch-clamp techniques and a mouse model of middle cerebral artery occlusion (MCAO)-induced focal ischemia were used. Surflex-Dock was used to dock the analogs into the pocket with default parameters.
RESULTS
Amiloride and its analogs inhibit ASIC1a currents expressed in Chinese hamster ovary cells with a potency rank order of benzamil > phenamil > 5-(N,N-dimethyl)amiloride (DMA) > amiloride > 5-(N,N-hexamethylene)amiloride (HMA) ≥ 5-(N-methyl-N-isopropyl)amiloride (MIA) > 5-(N-ethyl-N-isopropyl)amiloride (EIPA). In addition, amiloride and its analogs inhibit ASIC currents in cortical neurons with the same potency rank order. In mice, benzamil and EIPA decreased MCAO-induced infarct volume. Similar to its effect on the ASIC current, benzamil showed a much higher potency than EIPA.
CONCLUSION
Addition of a benzyl group to the terminal guanidinyl group resulted in enhanced inhibitory activity on ASIC1a. On the other hand, the bulky groups added to the 5-amino residues slightly decreased the activity. Among the tested amiloride analogs, benzamil is the most potent ASIC1a inhibitor.
Topics: Acid Sensing Ion Channels; Amiloride; Animals; Biophysics; CHO Cells; Cells, Cultured; Cerebral Cortex; Cricetinae; Cricetulus; Disease Models, Animal; Dose-Response Relationship, Drug; Electric Stimulation; Embryo, Mammalian; Infarction, Middle Cerebral Artery; Inhibitory Concentration 50; Male; Membrane Potentials; Mice; Mice, Inbred C57BL; Models, Molecular; Neurons; Patch-Clamp Techniques; Transfection
PubMed: 26890278
DOI: 10.1111/cns.12524 -
Biochimie Sep 1988Amiloride analogs inhibit a number of transmembrane Na+ transport systems: 1) the epithelium Na+ channel, 2) the Na+/H+ exchange system and 3) the Na+/Ca2+ exchange... (Review)
Review
Amiloride analogs inhibit a number of transmembrane Na+ transport systems: 1) the epithelium Na+ channel, 2) the Na+/H+ exchange system and 3) the Na+/Ca2+ exchange system. Structure--activity relationships using amiloride derivatives with selected modification of each of the functional groups of the molecule indicate that the 3 Na+ transporting systems have distinct pharmacological profiles. 5-N Disubstituted derivatives of amiloride, such as ethylisopropylamiloride are the most potent inhibitors of the Na+/H+ exchange system. Conversely, amiloride derivatives that are substituted on the guanidino moiety, such as phenamil, are potent inhibitors of the epithelium Na+ channel. It is thus possible, by using selected amiloride derivatives to inhibit selectively one or another of the Na+ transport systems.
Topics: Amiloride; Amphibians; Animals; Biological Transport; Carrier Proteins; Chickens; Epithelium; Kidney; Sodium Channels; Sodium-Calcium Exchanger; Sodium-Hydrogen Exchangers
PubMed: 2852509
DOI: 10.1016/0300-9084(88)90196-4 -
Biochemical and Biophysical Research... Dec 2023Sodium influx carried out by ion channels is one of the main regulators of water-salt and volume balance in cells of blood origin. Previously, we described...
Sodium influx carried out by ion channels is one of the main regulators of water-salt and volume balance in cells of blood origin. Previously, we described amiloride-insensitive ENaC-like channels in human myeloid leukemia K562 cells; the intracellular regulatory mechanisms of the channels are associated with actin cytoskeleton dynamics. Recently, an extracellular mechanism of ENaC-like channels activation in K562 cells by the action of serine protease trypsin has been revealed. The other extracellular pathways that modulate ENaC (epithelial Na channel) activity and sodium permeability in transformed blood cells are not yet fully investigated. Here, we study the action of capsazepine (CPZ), as δ-ENaC activator, on single channel activity in K562 cells in whole-cell patch clamp experiments. Addition of CPZ (2 μM) to the extracellular solution caused an activation of sodium channels with typical features; unitary conductance was 15.1 ± 0.8 pS. Amiloride derivative benzamil (50 μM) did not inhibit their activity. Unitary currents and conductance of CPZ-activated channels were higher in Na-containing extracellular solution than in Li, that is one of the main fingerprints of δ-ENaC. The results of RT-PCR analysis and immunofluorescence staining also confirmed the expression of δ-hENaC (as well as α-, β-, γ-ENaC) at the mRNA and protein level. These findings allow us to speculate that CPZ activates amiloride-insensitive ENaC-like channels that contain δ-ENaC in К562 cells. Our data reveal a novel extracellular mechanism for ENaC-like activation in human leukemia cells.
Topics: Humans; Amiloride; Epithelial Sodium Channels; Leukemia, Myeloid; Sodium; Oocytes
PubMed: 37944472
DOI: 10.1016/j.bbrc.2023.149187 -
Protein Science : a Publication of the... Oct 2023The SARS-CoV-2 envelope (E) protein forms a five-helix bundle in lipid bilayers whose cation-conducting activity is associated with the inflammatory response and...
The SARS-CoV-2 envelope (E) protein forms a five-helix bundle in lipid bilayers whose cation-conducting activity is associated with the inflammatory response and respiratory distress symptoms of COVID-19. E channel activity is inhibited by the drug 5-(N,N-hexamethylene) amiloride (HMA). However, the binding site of HMA in E has not been determined. Here we use solid-state NMR to measure distances between HMA and the E transmembrane domain (ETM) in lipid bilayers. C, N-labeled HMA is combined with fluorinated or C-labeled ETM. Conversely, fluorinated HMA is combined with C, N-labeled ETM. These orthogonal isotopic labeling patterns allow us to conduct dipolar recoupling NMR experiments to determine the HMA binding stoichiometry to ETM as well as HMA-protein distances. We find that HMA binds ETM with a stoichiometry of one drug per pentamer. Unexpectedly, the bound HMA is not centrally located within the channel pore, but lies on the lipid-facing surface in the middle of the TM domain. This result suggests that HMA may inhibit the E channel activity by interfering with the gating function of an aromatic network. These distance data are obtained under much lower drug concentrations than in previous chemical shift perturbation data, which showed the largest perturbation for N-terminal residues. This difference suggests that HMA has higher affinity for the protein-lipid interface than the channel pore. These results give insight into the inhibition mechanism of HMA for SARS-CoV-2 E.
Topics: Humans; Amiloride; SARS-CoV-2; Lipid Bilayers; COVID-19
PubMed: 37632140
DOI: 10.1002/pro.4755