Cyclic Nucleotide-gated Channels Research Articles

Membrane Lipid Nanodomains Modulate HCN Pacemaker Channels in Nociceptor DRG Neurons.

Cell membranes consist of heterogeneous lipid nanodomains that influence key cellular processes. Using FRET-based fluorescent assays and fluorescence lifetime imaging microscopy (FLIM), we found that the dimension of cholesterol-enriched ordered membrane domains (OMD) varies considerably, depending on specific cell types. Particularly, nociceptor dorsal root ganglion (DRG) neurons exhibit large OMDs. Disruption of OMDs potentiated action potential firing in nociceptor DRG neurons and facilitated the opening of native hyperpolarization-activated cyclic nucleotide-gated (HCN) pacemaker channels. This increased neuronal firing is partially due to an increased open probability and altered gating kinetics of HCN channels. The gating effect on HCN channels was likely due to a direct modulation of their voltage sensors by OMDs. In animal models of neuropathic pain, we observed reduced OMD size and a loss of HCN channel localization within OMDs. Additionally, cholesterol supplementation inhibited HCN channels and reduced neuronal hyperexcitability in pain models. These findings suggest that disturbances in lipid nanodomains play a critical role in regulating HCN channels within nociceptor DRG neurons, influencing pain modulation.

Genome-wide analysis of apple CNGC family allows the identification of MdCNGC15A negatively regulating apple salt tolerance

Calcium (Ca2+) is essential for signal conduction and plant growth. Cyclic nucleotide-gated channels (CNGCs) are Ca2+ transporters that regulate Ca2+ signalling and homeostasis by modulating its transmembrane transport, thereby influencing plant development as well as the biotic and abiotic stress responses. Although identified in numerous plant species, the CNGC family has not been characterized in apple until now. Here, 20 MdCNGCs were identified from the apple genome and were randomly distributed on 13 chromosomes. Phylogenetic analysis classified these MdCNGCs into five groups (I, Ⅱ, Ⅲ, Ⅳ-a, and Ⅳ-b), with five pairs of segmental duplicated genes being detected via collinearity analysis. Sequence alignment and analyses of gene structures, conserved motifs, and 3D structures indicated high structural conservation, particularly within groups. Yeast two-hybrid (Y2H) assays demonstrated interactions between most MdCNGCs and the Ca2+ receptor MdCaM7.1, except for MdCNGC1B and MdCNGC15A. Promoter analysis and expression profiling revealed significant responses to abiotic stress, particularly salt stress, in some MdCNGCs. Silencing MdCNGC15A significantly enhanced apple plants salt tolerance, while its overexpression in apple calli significantly decreased tolerance, as shown by transgenic analysis. Collectively, our results demonstrate the crucial role of MdCNGCs in abiotic stress responses and provide valuable insights for future functional and regulatory studies in apples.

Gene Therapy for Achromatopsia.

Achromatopsia is the most common cone dysfunction syndrome, affecting 1 in 30,000 people. It is an autosomal recessive disorder with a heterogeneous genetic background with variants reported in CNGA3, CNGB3, GNAT2, PDE6C, PDE6H, and ATF6. Up to 90% of achromatopsia patients harbour mutations in CNGA3 or CNB3, which encode for the alpha and beta subunits of the cone cyclic nucleotide-gated (CNG) channel in cone-specific phototransduction. The condition presents at birth or early infancy with poor visual acuity, nystagmus, photophobia, and colour vision loss in all axes. Multimodal retinal imaging has provided insightful information to characterise achromatopsia patients based on their genotype. There is no FDA-approved treatment for achromatopsia; however, studies have reported several preclinical gene therapies with anatomical and functional improvements reported in vivo. There are currently five gene therapy clinical trials registered for human patients at the phase I/II stage and for CNGA3 or CNGB3 causing achromatopsia. This review aims to discuss the genetics of achromatopsia, genotypic and phenotypic correlations in multimodal retinal imaging, and the developments and challenges in gene therapy clinical trials.

Transcriptome, hormonal, and secondary metabolite changes in leaves of DEFENSE NO DEATH 1 (DND1) silenced potato plants

DEFENSE NO DEATH 1 (DND1) is a cyclic nucleotide-gated ion channel protein. Earlier, it was shown that the silencing of DND1 in the potato (Solanum tuberosum L.) leads to resistance to late blight, powdery mildew, and gray mold diseases. At the same time, however, it can reduce plant growth and cause leaf necrosis. To obtain knowledge of the molecular events behind the pleiotropic effect of DND1 downregulation in the potato, metabolite and transcriptome analyses were performed on three DND1 silenced lines of the cultivar ‘Désirée.’ A massive increase in the salicylic acid content of leaves was detected. Concentrations of jasmonic acid and chlorogenic acid and their derivatives were also elevated. Expression of 1866 genes was altered in the same way in all three DND1 silenced lines, including those related to the synthesis of secondary metabolites. The activation of several alleles of leaf rust, late blight, and other disease resistance genes, as well as the induction of pathogenesis-related genes, was detected. WRKY and NAC transcription factor families were upregulated, whereas bHLHs were downregulated, indicating their central role in transcriptome changes. These results suggest that the maintenance of the constitutive defense state leads to the reduced growth of DND1 silenced potato plants.

Promotion of endoplasmic reticulum retrotranslocation by overexpression of E3 ubiquitin-protein ligase synoviolin 1 reduces endoplasmic reticulum stress and preserves cone photoreceptors in cyclic nucleotide-gated channel deficiency.

Cone photoreceptor cyclic nucleotide-gated (CNG) channels play an essential role in phototransduction and cellular Ca2+ homeostasis. Mutations in genes encoding the channel subunits CNGA3 and CNGB3 are associated with achromatopsia, progressive cone dystrophy, and early-onset macular degeneration. Cone loss in patients with achromatopsia and cone dystrophy associated with CNG channel mutations has been documented by optical coherence tomography and in mouse models of CNG channel deficiency. Cone death in CNG channel-deficient retinas involves endoplasmic reticulum (ER) stress-associated apoptosis, dysregulation of cellular/ER Ca2+ homeostasis, impaired protein folding/processing, and impaired ER-associated degradation (ERAD). The E3 ubiquitin-protein ligase synoviolin 1 (SYVN1) is the primary component of the SYVN1/SEL1L ER retrotranslocon responsible for ERAD. Previous studies have shown that manipulations that protect cones and reduce ER stress/cone death in CNG channel deficiency, such as increasing ER Ca2+ preservation or treatment with an ER chaperone, increase the expression of SYVN1 and other components of the ER retrotranslocon. The present work investigated the effects of SYVN1 overexpression. Intraocular injection of AAV5-IRBP/GNAT2-Syvn1 resulted in overexpression of SYVN1 in cones of CNG channel-deficient mice. Following treatment, cone density in Cnga3-/- mice was significantly increased, compared with untreated controls, outer segment localization of cone opsin was improved, and ER stress/apoptotic cell death was reduced. Overexpression of SYVN1 also led to increased expression levels of the retrotranslocon components, degradation in ER protein 1 (DERL1), ERAD E3 ligase adaptor subunit (SEL1L), and homocysteine inducible ER protein with ubiquitin-like domain 1 (HERPUD1). Moreover, overexpression of SYVN1 likely enhanced protein ubiquitination/proteasome degradation in CNG channel-deficient retinas. This study demonstrates the role of SYVN1/ERAD in cone preservation in CNG channel deficiency and supports the strategy of promoting ERAD for cone protection.

HCN channels are essential for the escape response of Paramecium.

When mechanical stimulation was applied to free swimming Paramecium, forward swimming velocity transiently increased due to activation of the posterior mechanosensory channels. The behavior response, known as "escape response," requires membrane hyperpolarization and the activation of K-channel type adenylate cyclases. Our hypothesis is that this escape response also involves activation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. HCN channels are activated by hyperpolarization and are modulated by cyclic nucleotides such as cAMP and cGMP. They play a critical role in many excitable cells in higher animals. If HCN channels act in Paramecium, this should help to enhance and prolong hyperpolarization, thereby increasing the swimming speed of Paramecium. This study used RNAi to examine the role of the HCN channel 1 in the escape responses by generating hcn1-gene knockdown cells (hcn1-KD). These cells showed reduced mechanically-stimulated escape responses and a lack of cGMP-dependent increases in swimming speed. Electrophysiological experiments demonstrated reduced hyperpolarization upon injection of large negative currents in hcn1-KD cells. This is consistent with a decrease in HCN1 channel activity and changes in the escape response. These findings suggest that HCN1 channels are K+ channels that regulate the escape response of Paramecium by amplifying the hyperpolarizations elicited by posterior mechanical stimulation.

Dual actions of chloroinconazide on pepper blight in Capsicum annuum: disruption of Phytophthora capsici mycelium and activation of CaCNGC9-mediated SA signaling.

Pepper blight, caused by Phytophthora capsici, is a devastating disease that seriously threatens pepper production worldwide. With the emergence of resistance in P. capsici against conventional fungicides, there is an urgent need to explore novel alternatives for pepper blight management. This study aims to assess the inhibitory effect of chloroinconazide (CHI), a compound synthesized from tryptophan, against pepper blight, and to explore its potential mechanisms of action. The results demonstrated that CHI effectively targeted P. capsici, disrupting its growth and mycelial structure, which resulted in the release of dissolved intracellular substances. Additionally, CHI significantly inhibited the sporangium formation, zoospores release, and zoospores germination, thereby reducing the re-infection of P. capsici. In contrast, the commercial pesticide methylaxyl only inhibited mycelial growth and had limited effect on re-infection, while azoxystrobin inhibited re-infection but had a weak inhibitory effect on mycelial growth. Furthermore, CHI activated the salicylic acid (SA) signaling pathway-mediated immune response to inhibit P. capsici infection in pepper, with this activation being contingent upon cyclic nucleotide-gated ion channel CaCNGC9. CHI exhibited potent dual inhibitory effects on P. capsici by disrupting mycelial structure and activating the CaCNGC9-mediated SA signaling pathway. These dual mechanisms of action suggested that CHI could serve as a promising alternative chemical fungicide for the effective management of pepper blight, offering a new approach to control this devastating disease. Our findings highlighted the potential of CHI as a sustainable and efficient solution to combat the increasing resistance of P. capsici to conventional fungicides, ensuring better crop protection and yield. © 2024 Society of Chemical Industry.

Drosophila HCN mediates gustatory homeostasis by preserving sensillar transepithelial potential in sweet environments.

Establishing transepithelial ion disparities is crucial for sensory functions in animals. In insect sensory organs called sensilla, a transepithelial potential, known as the sensillum potential (SP), arises through active ion transport across accessory cells, sensitizing receptor neurons such as mechanoreceptors and chemoreceptors. Because multiple receptor neurons are often co-housed in a sensillum and share SP, niche-prevalent overstimulation of single sensory neurons can compromise neighboring receptors by depleting SP. However, how such potential depletion is prevented to maintain sensory homeostasis remains unknown. Here, we find that the Ih-encoded hyperpolarization-activated cyclic nucleotide-gated (HCN) channel bolsters the activity of bitter-sensing gustatory receptor neurons (bGRNs), albeit acting in sweet-sensing GRNs (sGRNs). For this task, HCN maintains SP despite prolonged sGRN stimulation induced by the diet mimicking their sweet feeding niche, such as overripe fruit. We present evidence that Ih-dependent demarcation of sGRN excitability is implemented to throttle SP consumption, which may have facilitated adaptation to a sweetness-dominated environment. Thus, HCN expressed in sGRNs serves as a key component of a simple yet versatile peripheral coding that regulates bitterness for optimal food intake in two contrasting ways: sweet-resilient preservation of bitter aversion and the previously reported sweet-dependent suppression of bitter taste.

Dopaminergic amacrine cells express HCN channels in the developing and adult mouse retina.

To determine the molecular and functional expression of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in developing and mature dopaminergic amacrine cells (DACs), the sole source of ocular dopamine that plays a vital role in visual function and eye development. HCN channels are encoded by isoforms 1-4. HCN1, HCN2, and HCN4 were immunostained in retinal slices obtained from mice at postnatal day 4 (P4), P8, and P12 as well as in adults. Each HCN channel isoform was also immunostained with tyrosine hydroxylase, a marker for DACs, at P12 and adult retinas. Genetically-marked DACs were recorded in flat-mount retina preparation using a whole-cell current-clamp technique. HCN1 was expressed in rods/cones, amacrine cells, and retinal ganglion cells (RGCs) at P4, along with bipolar cells by P12. Different from HCN1, HCN2 and HCN4 were each expressed in amacrine cells and RGCs at P4, along with bipolar cells by P8, and in rods/cones by P12. Double immunostaining shows that each of the three isoforms was expressed in approximately half of DACs at P12 but in almost all DACs in adults. Electrophysiology results demonstrate that HCN channel isoforms form functional HCN channels, and the pharmacological blockade of HCN channels reduced the spontaneous firing frequency in most DACs. Each class of retinal neurons may use different isoforms of HCN channels to function during development. HCN1, HCN2, and HCN4 form functional HCN channels in DACs, which appears to modulate their spontaneous firing activity.

Melatonin-mediated low-temperature tolerance of cucumber seedlings requires Ca2+/CPKs signaling pathway

Melatonin (Mel) is recognized as a prominent plant growth regulator. This study investigated the alleviating effect of Mel pretreatment on growth inhibition caused by low-temperature (LT) stress (10 °C/6 °C) in cucumber seedlings and explored the role of the Ca2+/Calcium-dependent protein kinases (CPKs) signaling pathway in Mel-regulated LT tolerance. The main results are as follows: compared to LT treatment alone, 100 μM Mel increased both the content of Ca2+ (highest about 42.01%) and the expression levels of Ca2+ transporter and cyclic nucleotide-gated channel (CNGC) genes under LT. Similarly, Mel enhanced the content of CPKs (highest about 27.49%) and the expression levels of CPKs family genes in cucumber leaves under LT. Additionally, pretreatment with 100 μM Mel for three days strengthened the antioxidant defense and photosynthesis of seedlings under LT. Genes in the ICE-CBF-COR pathway and the MAPK cascade were upregulated by Mel, with maximum upregulations reaching approximately 2.5-fold and 1.9-fold, respectively, thus conferring LT tolerance to cucumber seedlings. However, the above beneficial effects of Mel were weakened by co-treatment with calcium signaling blockers (LaCl3 or EGTA) or CPKs inhibitors (TFP or W-7), suggesting that the Ca2+/CPKs pathway is involved in the Mel-mediated regulation of LT tolerance. In conclusion, this study revealed that Mel can alleviate growth inhibition in cucumber seedlings under LT stress and demonstrated that the Ca2+/CPKs signaling pathway is crucial for the Mel-mediated enhancement of LT tolerance. The findings hold promise for providing theoretical insights into the application of Mel in agricultural production and for investigating its underlying mechanisms of action.

Tau-mediated synaptic dysfunction is coupled with HCN channelopathy.

In tauopathies, altered tau processing correlates with impairments in synaptic density and function. Changes in hyperpolarization-activated cyclic nucleotide-gated (HCN) channels contribute to disease-associated abnormalities in multiple neurodegenerative diseases. To investigate the link between tau and HCN channels, we performed histological, biochemical, ultrastructural, and functional analyses of hippocampal tissues from Alzheimer's disease (AD), age-matched controls, Tau35 mice, and/or Tau35 primary hippocampal neurons. Expression of specific HCN channels is elevated in post mortem AD hippocampus. Tau35 mice develop progressive abnormalities including increased phosphorylated tau, enhanced HCN channel expression, decreased dendritic branching, reduced synapse density, and vesicle clustering defects. Tau35 primary neurons show increased HCN channel expression enhanced hyperpolarization-induced membrane voltage "sag" and changes in the frequency and kinetics of spontaneous excitatory postsynaptic currents. Our findings are consistent with a model in which pathological changes in tauopathies impact HCN channels to drive network-wide structural and functional synaptic deficits. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are functionally linked to the development of tauopathy. Expression of specific HCN channels is elevated in the hippocampus in Alzheimer's disease and the Tau35 mouse model of tauopathy. Increased expression of HCN channels in Tau35 mice is accompanied by hyperpolarization-induced membrane voltage "sag" demonstrating a detrimental effect of tau abnormalities on HCN channel function. Tau35 expression alters synaptic organization, causing a loosened vesicle clustering phenotype in Tau35 mice.

Cellular and circuit architecture of the lateral septum for reward processing

The lateral septum (LS) is composed of heterogeneous cell types that are important for various motivated behaviors. However, the transcriptional profiles, spatial arrangement, function, and connectivity of these cell types have not been systematically studied. Using single-nucleus RNA sequencing, we delineated diverse genetically defined cell types in the LS that play distinct roles in reward processing. Notably, we found that estrogen receptor 1 (Esr1)-expressing neurons in the ventral LS (LSEsr1) are key drivers of reward seeking via projections to the ventral tegmental area, and these neurons play an essential role in methamphetamine (METH) reward and METH-seeking behavior. Extended exposure to METH increases the excitability of LSEsr1 neurons by upregulating hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, thereby contributing to METH-induced locomotor sensitization. These insights not only elucidate the intricate molecular, circuit, and functional architecture of the septal region in reward processing but also reveal a neural pathway critical for METH reward and behavioral sensitization.

HCN channels in the lateral habenula regulate pain and comorbid depressive-like behaviors in mice.

Comorbid anxiodepressive-like symptoms (CADS) in chronic pain are closely related to the overactivation of the lateral habenula (LHb). Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels have been implicated to play a key role in regulating neuronal excitability. However, the role of HCN channels in the LHb during CADS has not yet been characterized. This study aimed to investigate the effect of HCN channels in the LHb on CADS during chronic pain. After chronic neuropathic pain induction by spared nerve injury (SNI), mice underwent a sucrose preference test, forced swimming test, tail suspension test, open-field test, and elevated plus maze test to evaluate their anxiodepressive-like behaviors. Electrophysiological recordings, immunohistochemistry, Western blotting, pharmacological experiments, and virus knockdown strategies were used to investigate the underlying mechanisms. Evident anxiodepressive-like behaviors were observed 6w after the SNI surgery, accompanied by increased neuronal excitability, enhanced HCN channel function, and increased expression of HCN2 isoforms in the LHb. Either pharmacological inhibition or virus knockdown of HCN2 channels significantly reduced LHb neuronal excitability and ameliorated both pain and depressive-like behaviors. Our results indicated that the LHb neurons were hyperactive under CADS in chronic pain, and this hyperactivation possibly resulted from the enhanced function of HCN channels and up-regulation of HCN2 isoforms.

Ionic mechanisms involved in arginine vasopressin-mediated excitation of auditory cortical and thalamic neurons

The axons containing arginine vasopressin (AVP) from the hypothalamus innervate a variety of structures including the cerebral cortex, thalamus, hippocampus and amygdala. A plethora amount of evidence indicates that activation of the V1a subtype of the vasopressin receptors facilitates anxiety-like and fear responses. As an essential structure involved in fear and anxiety responses, the amygdala, especially the lateral nucleus of amygdala (LA), receives glutamatergic innervations from the auditory cortex and auditory thalamus where high density of V1a receptors have been detected. However, the roles and mechanisms of AVP in these two important areas have not been determined, which prevents the understanding of the mechanisms whereby V1a activation augments anxiety and fear responses. Here, we used coronal brain slices and studied the effects of AVP on neuronal activities of the auditory cortical and thalamic neurons. Our results indicate that activation of V1a receptors excited both auditory cortical and thalamic neurons. In the auditory cortical neurons, AVP increased neuronal excitability by depressing multiple subtypes of inwardly rectifying K+ (Kir) channels including the Kir2 subfamily, the ATP-sensitive K+ channels and the G protein-gated inwardly rectifying K+ (GIRK) channels, whereas activation of V1a receptors excited the auditory thalamic neurons by depressing the Kir2 subfamily of the Kir channels as well as activating the hyperpolarization-activated cyclic nucleotide-gated (HCN) channels and a persistent Na+ channel. Our results may help explain the roles of V1a receptors in facilitating fear and anxiety responses.Categories: Cell Physiology.

Inhibition of a cyclic nucleotide-gated channel on neuronal cilia activates unfolded protein response in intestinal cells to promote longevity

Ciliary defects are linked to ciliopathies, but impairments in the sensory cilia of Caenorhabditis elegans neurons extend lifespan, a phenomenon with previously unclear mechanisms. Our study reveals that neuronal cilia defects trigger the unfolded protein response of the endoplasmic reticulum (UPR ER ) within intestinal cells, a process dependent on the insulin/insulin-like growth factor 1 (IGF-1) signaling transcription factor and the release of neuronal signaling molecules. While inhibiting UPR ER doesn’t alter the lifespan of wild-type worms, it normalizes the extended lifespan of ciliary mutants. Notably, deactivating the cyclic nucleotide-gated (CNG) channel TAX-4 on the ciliary membrane promotes lifespan extension through a UPR ER -dependent mechanism. Conversely, constitutive activation of TAX-4 attenuates intestinal UPR ER in ciliary mutants. Administering a CNG channel blocker to worm larvae activates intestinal UPR ER and increases adult longevity. These findings suggest that ciliary dysfunction in sensory neurons triggers intestinal UPR ER , contributing to lifespan extension and implying that transiently inhibiting ciliary channel activity may effectively prolong lifespan.

Glucagon-like peptide-1 increases heart rate by a direct action on the sinus node.

Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) are increasingly used to treat type 2 diabetes and obesity. Albeit cardiovascular outcomes generally improve, treatment with GLP-1 RAs is associated with increased heart rate, the mechanism of which is unclear. We employed a large animal model, the female landrace pig, and used multiple in vivo and ex vivo approaches including pharmacological challenges, electrophysiology, and high-resolution mass spectrometry to explore how GLP-1 elicits an increase in heart rate. In anaesthetized pigs, neither cervical vagotomy, adrenergic blockers (alpha, beta, or combined alpha-beta blockade), ganglionic blockade (hexamethonium), nor inhibition of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels (ivabradine) abolished the marked chronotropic effect of GLP-1. GLP-1 administration to isolated perfused pig hearts also increased heart rate, which was abolished by GLP-1 receptor blockade. Electrophysiological characterization of GLP-1 effects in vivo and in isolated perfused hearts localized electrical modulation to the atria and conduction system. In isolated sinus nodes, GLP-1 administration shortened the action potential cycle length of pacemaker cells and shifted the site of earliest activation. The effect was independent of HCN blockade. Collectively, these data support a direct effect of GLP-1 on GLP-1 receptors within the heart. Consistently, single nucleus RNA sequencing showed GLP-1 receptor expression in porcine pacemaker cells. Quantitative phosphoproteomics analyses of sinus node samples revealed that GLP-1 administration leads to phosphorylation changes of calcium cycling proteins of the sarcoplasmic reticulum, known to regulate heart rate. GLP-1 has direct chronotropic effects on the heart mediated by GLP-1 receptors in pacemaker cells of the sinus node, inducing changes in action potential morphology and the leading pacemaker site through a calcium signalling response characterized by PKA-dependent phosphorylation of Ca2+ cycling proteins involved in pacemaking. Targeting the pacemaker calcium clock may be a strategy to lower heart rate in people treated with GLP-1 RAs.

High-Resolution Proteomics Unravel a Native Functional Complex of Cav1.3, SK3, and Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels in Midbrain Dopaminergic Neurons.

Pacemaking activity in substantia nigra dopaminergic neurons is generated by the coordinated activity of a variety of distinct somatodendritic voltage- and calcium-gated ion channels. We investigated whether these functional interactions could arise from a common localization in macromolecular complexes where physical proximity would allow for efficient interaction and co-regulations. For that purpose, we immunopurified six ion channel proteins involved in substantia nigra neuron autonomous firing to identify their molecular interactions. The ion channels chosen as bait were Cav1.2, Cav1.3, HCN2, HCN4, Kv4.3, and SK3 channel proteins, and the methods chosen to determine interactions were co-immunoprecipitation analyzed through immunoblot and mass spectrometry as well as proximity ligation assay. A macromolecular complex composed of Cav1.3, HCN, and SK3 channels was unraveled. In addition, novel potential interactions between SK3 channels and sclerosis tuberous complex (Tsc) proteins, inhibitors of mTOR, and between HCN4 channels and the pro-degenerative protein Sarm1 were uncovered. In order to demonstrate the presence of these molecular interactions in situ, we used proximity ligation assay (PLA) imaging on midbrain slices containing the substantia nigra, and we could ascertain the presence of these protein complexes specifically in substantia nigra dopaminergic neurons. Based on the complementary functional role of the ion channels in the macromolecular complex identified, these results suggest that such tight interactions could partly underly the robustness of pacemaking in dopaminergic neurons.

Pacemaker Channels and the Chronotropic Response in Health and Disease

Loss or dysregulation of the normally precise control of heart rate via the autonomic nervous system plays a critical role during the development and progression of cardiovascular disease—including ischemic heart disease, heart failure, and arrhythmias. While the clinical significance of regulating changes in heart rate, known as the chronotropic effect, is undeniable, the mechanisms controlling these changes remain not fully understood. Heart rate acceleration and deceleration are mediated by increasing or decreasing the spontaneous firing rate of pacemaker cells in the sinoatrial node. During the transition from rest to activity, sympathetic neurons stimulate these cells by activating β-adrenergic receptors and increasing intracellular cyclic adenosine monophosphate. The same signal transduction pathway is targeted by positive chronotropic drugs such as norepinephrine and dobutamine, which are used in the treatment of cardiogenic shock and severe heart failure. The cyclic adenosine monophosphate–sensitive hyperpolarization–activated current (I f ) in pacemaker cells is passed by hyperpolarization-activated cyclic nucleotide-gated cation channels and is critical for generating the autonomous heartbeat. In addition, this current has been suggested to play a central role in the chronotropic effect. Recent studies demonstrate that cyclic adenosine monophosphate–dependent regulation of HCN4 (hyperpolarization-activated cyclic nucleotide-gated cation channel isoform 4) acts to stabilize the heart rate, particularly during rapid rate transitions induced by the autonomic nervous system. The mechanism is based on creating a balance between firing and recently discovered nonfiring pacemaker cells in the sinoatrial node. In this way, hyperpolarization-activated cyclic nucleotide-gated cation channels may protect the heart from sinoatrial node dysfunction, secondary arrhythmia of the atria, and potentially fatal tachyarrhythmia of the ventricles. Here, we review the latest findings on sinoatrial node automaticity and discuss the physiological and pathophysiological role of HCN pacemaker channels in the chronotropic response and beyond.

A cyclic nucleotide-gated channel gene HcCNGC21 positively regulates salt and drought stress responses in kenaf (Hibiscus cannabinus L.)

Cyclic Nucleotide-Gated Channels (CNGCs) serve as Ca2+ permeable cation transport pathways, which are involved in the regulation of various biological functions such as plant cell ion selective permeability, growth and development, responses to biotic and abiotic stresses. At the present study, a total of 31 CNGC genes were identified and bioinformatically analyzed in kenaf. Among these genes, HcCNGC21 characterized to localize at the plasma membrane, with the highest expression levels in leaves, followed by roots. In addition, HcCNGC21 could be significantly induced under salt or drought stress. Virus-induced gene silencing (VIGS) of HcCNGC21 in kenaf caused notable growth inhibition under salt or drought stress, characterized by reductions in plant height, stem diameter, leaf area, root length, root surface area, and root tip number. Meanwhile, the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) were significantly decreased, accompanied by reduced levels of osmoregulatory substances and total chlorophyll content. However, ROS accumulation and Na+ content increased. The expression of stress-responsive genes, such as HcSOD, HcPOD, HcCAT, HcERF3, HcNAC29, HcP5CS, HcLTP, and HcNCED, was significantly downregulated in these silenced lines. However, under salt or drought stress, the physiological performance and expression of stress-related genes in transgenic Arabidopsis thaliana plants overexpressing HcCNGC21 were diametrically opposite to those of TRV2-HcCNGC21 kenaf line. Yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays revealed that HcCNGC21 interacts with HcAnnexin D1. These findings collectively underscore the positive role of HcCNGC21 in plant resistance to salt and drought stress.

Genotypic and Phenotypic Characterization of a Cohort of Patients Affected by Rod Cyclic Nucleotide Channel-Associated Retinitis Pigmentosa.

Retinitis pigmentosa (RP), a heterogeneous inherited retinal disorder causing gradual vision loss, affects over 1 million people worldwide. Pathogenic variants in CNGA1 and CNGB1 genes, respectively, accounting for 1% and 4% of cases, impact the cyclic nucleotide-gated channel in rod photoreceptor cells. The aim of this study was to describe and compare genotypic and clinical characteristics of a cohort of patients with CNGA1- or CNGB1-related RP and to explore potential genotype-phenotype correlations. The following data from patients with CNGA1- or CNGB1-related RP, followed in five Italian inherited retinal degenerations services, were retrospectively collected: genetic variants in CNGA1 and CNGB1, best-corrected visual acuity (BCVA), ellipsoid zone (EZ) width, fundus photographs, and short-wavelength fundus autofluorescence (SW-AF) images. Comparisons and correlation analyses were performed by first dividing the cohort in two groups according to the gene responsible for the disease (CNGA1 and CNGB1 groups). In parallel, the whole cohort of RP patients was divided into two other groups, according to the expected impact of the variants at protein level (low and high group). In total, 29 patients were recruited, 11 with CNGA1- and 18 with CNGB1-related RP. In both CNGA1 and CNGB1, 5 novel variants in CNGA1 and 5 in CNGB1 were found. BCVA was comparable between CNGA1 and CNGB1 groups, as well as between low and high groups. CNGA1 group had a larger mean EZ width compared to CNGB1 group, albeit not statistically significant, while EZ width did not differ between low and high groups A statistically significant correlation between EZ width and BCVA as well as between EZ width and age were observed in the whole cohort of RP patients. Fundus photographs of all patients in the cohort showed classic RP pattern, and in SW-AF images an hyperautofluorescent ring was observed in 14/21 patients. Rod CNG channel-associated RP was demonstrated to be a slowly progressive disease in both CNGA1- and CNGB1-related forms, making it an ideal candidate for gene augmentation therapies.