Patch clamp nAChR

UNLABELLED: Cochlear outer hair cells (OHCs) are electromotile and are implicated in mechanisms of amplification of responses to sound that enhance sound sensitivity and frequency tuning. They send information to the brain through glutamatergic synapses onto a small subpopulation of neurons of the ascending auditory nerve, the type II spiral ganglion neurons (SGNs). The OHC synapses onto type II SGNs are sparse and weak, suggesting that type II SGNs respond primarily to loud and possibly damaging levels of sound...

We describe molecular-level functional changes in the α4β2 nicotinic acetylcholine receptor (nAChR) by a leucine residue insertion in the M2 transmembrane domain of the α4 subunit associated with sleep-related hyperkinetic epilepsy. Measurements of agonist-elicited single channel currents reveal the primary effect is to stabilize the open channel state while the secondary effect is to promote reopening of the channel. These dual effects prolong the durations of bursts of channel openings equally for the two major stoichiometric forms of the receptor, (α4)2 (β2)3 and (α4)3 (β2)2 , indicating the functional impact is independent of mutant copy number per receptor...

BACKGROUND AND PURPOSE: α4β2 nicotinic acetylcholine (nACh) receptors assemble in two stoichiometric forms, one of which is potentiated by calcium. The sites of calcium binding that underpin potentiation are not known. EXPERIMENTAL APPROACH: To identify calcium binding sites, we applied cryo-electron microscopy (cryo-EM) and molecular dynamics (MD) simulations to each stoichiometric form of the α4β2 nACh receptor in the presence of calcium ions...

Insect neuronal nicotinic acetylcholine receptors (nAChRs) are transmembrane receptors that play a key role in the development and synaptic plasticity of both vertebrates and invertebrates and are considered to be major targets of neonicotinoid insecticides. We used dorsal unpaired median (DUM) neurons, which are insect neurosecretory cells, in order to explore the intracellular mechanisms leading to the regulation of insect neuronal nAChRs in more detail. Using whole-cell patch-clamp and fura-2AM calcium imaging techniques, we found that a novel CaMKK/AMPK pathway could be involved in the intracellular regulation of DUM neuron nAChRs...

Tinnitus impacts between 10-20% of the population. Individuals most troubled by their tinnitus have their attention bound to and are distracted by, their tinnitus percept. While numerous treatments to ameliorate tinnitus have been tried, no therapeutic approach has been clinically accepted. The present study used an established condition-suppression noise-exposure rat model of tinnitus to: (1) examine tinnitus-related changes in nAChR function of layer 5 pyramidal (PNs) and of vasoactive intestinal peptide (VIP) neurons in primary auditory cortex (A1) and (2) examine how the partial desensitizing nAChR agonists, sazetidine-A and varenicline, can act as potential therapeutic agents in the treatment of tinnitus...

Although nicotinic acetylcholine receptors (nAChRs) are commonly associated with neurons in the brain and periphery, recent data indicate they are also expressed in non-neuronal tissues. We recently found the alpha7 (α7nAChR) subunit is highly expressed in human airway smooth muscle (hASM) with substantial increase in asthmatics, but their functionality remains unknown. We investigated the location and functional role of α7nAChRs in hASM cells from normal vs. mild-moderate asthmatic patients. Immunostaining and protein analyses showed α7nAChR in the plasma membrane including in asthmatics...

OBJECTIVE: The pro-inflammatory cytokine interleukin-1β (IL-1β) plays a central role in host defense against infections. High systemic IL-1β levels, however, promote the pathogenesis of inflammatory disorders. Therefore, mechanisms controlling IL-1β release are of substantial clinical interest. Recently, we identified a cholinergic mechanism inhibiting the ATP-mediated IL-1β release by human monocytes via nicotinic acetylcholine receptor (nAChR) subunits α7, α9 and/or α10...

BACKGROUND AND PURPOSE: Local blood flow regulation relies mostly on the coordination between neurons and pericyte-containing capillaries. Pericyte relaxation and contraction are influenced by various vasoactive substances and regulated by neurotransmitters. α7 nicotinic acetylcholine receptors (α7-nAChRs), involved in the regulation of vascular function and inhibitory GABA systems, have neuroprotective effects against central nervous system diseases. Although α7-nAChRs are found throughout the retina, their contribution to the retinal capillary tone remains unknown...

Ladybird beetles (Coleoptera: Coccinellidae) possess strong chemical defences that are secreted in response to stress and are also found on the coating of eggs, which are rich in alkaloids that are responsible for their toxicity to other species. Recent studies have shown that alkaloids from several species of ladybird beetle can target nicotinic acetylcholine receptors (nAChRs) acting as receptor antagonists. Here, we have explored the actions of (-)-adaline, found in the 2-spot ( Adalia bipunctata ) and 10-spot ( Adalia decempunctata ) ladybirds, on both mammalian (α1β1γδ, α7, α4β2, α3β4) and insect nAChRs using patch-clamp of TE671 cells and locust brain neurons natively expressing nAChRs, as well as two-electrode voltage clamp of Xenopus laevis oocytes recombinantly expressing nAChRs...

We describe genetic and molecular-level functional alterations in the α4β2 neuronal nicotinic acetylcholine receptor (nAChR) from a patient with sleep-related hyperkinetic epilepsy and a family history of epilepsy. Genetic sequencing revealed a heterozygous variant c.851C>G in the CHRNA4 gene encoding the α4 subunit, resulting in the missense mutation p.Ser284Trp. Patch clamp recordings from genetically engineered nAChRs incorporating the α4-Ser284Trp subunit revealed aberrant channel openings in the absence of agonist and markedly prolonged openings in its presence...

Cannabidiol (CBD), an important terpenoid compound from marijuana with no psychoactive effects, has become of great pharmaceutical interest for several health conditions. As CBD is a multitarget drug, there is a need to establish the molecular mechanisms by which CBD may exert therapeutic as well as adverse effects. The α7 nicotinic acetylcholine receptor (α7 nAChR) is a cation-permeable ACh-gated channel present in the nervous system and in non-neuronal cells. It is involved in different pathological conditions, including neurological and neurodegenerative disorders, inflammation, and cancer...

Noopept (NP) is a proline-containing dipeptide with nootropic and neuroprotective properties. We have previously shown that NP significantly increased the frequency of spontaneous IPSCs in hippocampal CA1 pyramidal cells mediated by the activation of inhibitory interneurons in stratum radiatum. The cholinergic system plays an important role in the performance of cognitive functions, furthermore multiple behavioral and clinical facts link NP with the cholinergic system. The present study was undertaken to reveal the possible interaction of NP with neuronal nicotinic acetylcholine receptors (nAChRs)...

Apamin is often cited as one of the few substances selectively acting on small-conductance Ca2+ -activated potassium channels (KCa 2). However, published pharmacological and structural data remain controversial. Here, we investigated the molecular pharmacology of apamin by two-electrode voltage-clamp in Xenopus laevis oocytes and patch-clamp in HEK293, COS7, and CHO cells expressing the studied ion channels, as well as in isolated rat brain neurons. The microtitre broth dilution method was used for antimicrobial activity screening...

Nicotinic acetylcholine receptors (nAChRs) are involved in a great range of physiological and pathological conditions. Since they are transmembrane proteins, they interact strongly with the lipids surrounding them. Thus, the plasma membrane composition and heterogeneity play an essential role for the correct nAChR function, on the one hand, and the nAChR influences its immediate lipid environment, on the other hand. The aim of this work was to investigate in more detail the role of the biophysical properties of the membrane in nAChR function and vice versa, focusing on the relationship between Chol and nAChRs...

The hippocampus is an important area for memory encoding and retrieval and is the location of spike timing-dependent plasticity (STDP), a basic phenomenon of learning and memory. STDP is facilitated if acetylcholine (ACh) is released from cholinergic neurons during attentional processes. However, it is unclear how ACh influences postsynaptic changes during STDP induction and determines the STDP magnitude. To address these issues, we obtained patch clamp recordings from CA1 pyramidal neurons to evaluate the postsynaptic changes during stimuli injection in Schaffer collaterals by quantifying baseline amplitudes (i...

The α7 nicotinic acetylcholine receptor (nAChR) is present in neuronal and non-neuronal cells and has anti-inflammatory actions. Molecular dynamics simulations suggested that α7 nAChR interacts with a region of the SARS-CoV-2 spike protein (S), and a potential contribution of nAChRs to COVID-19 pathophysiology has been proposed. We applied whole-cell and single-channel recordings to determine whether a peptide corresponding to the Y674-R685 region of the S protein can directly affect α7 nAChR function. The S fragment exerts a dual effect on α7...

There are about twice as many women as men who experience depression during their lifetime. Although life circumstances and especially exposure to stressful situations constitute a major risk factor to develop depression, the underlying mechanisms have yet to be unraveled. We employed the chronic social defeat procedure to elicit depressive-like symptoms in females and ketamine to validate the model. We performed ex-vivo patch clamp recordings to assess cellular adaptations and used pharmacological agents to dissect these deregulations...

The physical interaction and functional cross talk among the different subtypes of neuronal nicotinic acetylcholine receptors (nAChRs) expressed in the various tissues is unknown. Here, we have investigated this issue between the only two nAChRs subtypes expressed, the α7 and α3β4 subtypes, in a human native neuroendocrine cell (the chromaffin cell) using electrophysiological patch-clamp, fluorescence, and Förster resonance energy transfer (FRET) techniques. Our data show that α7 and α3β4 receptor subtypes require their mutual and maximal efficacy of activation to increase their expression, to avoid their desensitization, and therefore, to increase their activity...

Retinal bipolar cells are second-order neurons that transmit basic features of the visual scene to postsynaptic partners. However, their contribution to motion detection has not been fully appreciated. Here, we demonstrate that cholinergic feedback from starburst amacrine cells (SACs) to certain presynaptic bipolar cells via alpha-7 nicotinic acetylcholine receptors (α7-nAChRs) promotes direction-selective signaling. Patch clamp recordings reveal that distinct bipolar cell types making synapses at proximal SAC dendrites also express α7-nAChRs, producing directionally skewed excitatory inputs...

Alterations in the polyamine and amino acid (tyrosine) moieties of philanthotoxin-343 (PhTX-343) were investigated for their effects on the antagonism of nicotinic acetylcholine receptors (nAChRs) isolated from the locust ( Schistocerca gregaria ) mushroom body. Through whole-cell patch-clamp recordings, the philanthotoxin analogues in this study were shown to cause inhibition of the inward current when co-applied with acetylcholine (ACh). PhTX-343 (IC50 = 0.80 μM at -75 mV) antagonised locust nAChRs in a use-dependent manner, suggesting that it acts as an open-channel blocker...