The Concise Guide to PHARMACOLOGY 2023/24: Ion channels.

The Concise Guide to PHARMACOLOGY 2023/24 is the sixth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of approximately 1800 drug targets, and over 6000 interactions with about 3900 ligands. There is an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (https://www.guidetopharmacology.org/), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes almost 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.16178. Ion channels are one of the six major pharmacological targets into which the Guide is divided, with the others being: G protein-coupled receptors, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2023, and supersedes data presented in the 2021/22, 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

mRNA trans-splicing dual AAV vectors for (epi)genome editing and gene therapy.

Large genes including several CRISPR-Cas modules like gene activators (CRISPRa) require dual adeno-associated viral (AAV) vectors for an efficient in vivo delivery and expression. Current dual AAV vector approaches have important limitations, e.g., low reconstitution efficiency, production of alien proteins, or low flexibility in split site selection. Here, we present a dual AAV vector technology based on reconstitution via mRNA trans-splicing (REVeRT). REVeRT is flexible in split site selection and can efficiently reconstitute different split genes in numerous in vitro models, in human organoids, and in vivo. Furthermore, REVeRT can functionally reconstitute a CRISPRa module targeting genes in various mouse tissues and organs in single or multiplexed approaches upon different routes of administration. Finally, REVeRT enabled the reconstitution of full-length ABCA4 after intravitreal injection in a mouse model of Stargardt disease. Due to its flexibility and efficiency REVeRT harbors great potential for basic research and clinical applications.

Preliminary Assessment of Pre-Electroconvulsive Therapy Evaluation Practices in European Countries: The Need for Guidelines.

OBJECTIVES: Pre-electroconvulsive therapy (ECT) evaluation is an essential part of ECT preparation, a standard treatment in the psychiatric field. However, no routine pre-ECT evaluation has been published so far. This preliminary study aimed to explore different practices in pre-ECT evaluation across European countries. METHODS: The data were collected as a snowball sample approach using an online survey from September 2019 to April 2020. The final analysis included data from 18 clinics placed in 16 European countries. RESULTS: Regulations on the pre-ECT evaluation were found in 9 countries. All clinics reported doing complete blood count, serum electrolytes, and renal function analysis as a part of regular laboratory testing, alongside with a cardiovascular assessment. Ten clinics reported using psychiatric scales. Six clinics reported doing a cognitive assessment, of which all had regulations on the pre-ECT evaluation. Not one evaluation had the same sets of procedures and diagnostics. CONCLUSIONS: The differences in assessment approaches mirror high variability of the pre-ECT evaluation practice across Europe. Cognitive assessment and objectification of psychiatric symptoms should be a regular part of the pre-ECT evaluation because of the monitoring of the most common adverse effect and observing the clinical response to ECT. Standardization of the pre-ECT evaluation and ECT in general would remove criticisms and opposition to the treatment, make it based on the best of our knowledge, and provide a method respectful of patients' best interests and rights.

dCas9-VPR-mediated transcriptional activation of functionally equivalent genes for gene therapy.

Many disease-causing genes possess functionally equivalent counterparts, which are often expressed in distinct cell types. An attractive gene therapy approach for inherited disorders caused by mutations in such genes is to transcriptionally activate the appropriate counterpart(s) to compensate for the missing gene function. This approach offers key advantages over conventional gene therapies because it is mutation- and gene size-independent. Here, we describe a protocol for the design, execution and evaluation of such gene therapies using dCas9-VPR. We offer guidelines on how to identify functionally equivalent genes, design and clone single guide RNAs and evaluate transcriptional activation in vitro. Moreover, focusing on inherited retinal diseases, we provide a detailed protocol on how to apply this strategy in mice using dual recombinant adeno-associated virus vectors and how to evaluate its functionality and off-target effects in the target tissue. This strategy is in principle applicable to all organisms that possess functionally equivalent genes suitable for transcriptional activation and addresses pivotal unmet needs in gene therapy with high translational potential. The protocol can be completed in 15-20 weeks.

Maybe you can turn me on: CRISPRa-based strategies for therapeutic applications.

Since the revolutionary discovery of the CRISPR-Cas technology for programmable genome editing, its range of applications has been extended by multiple biotechnological tools that go far beyond its original function as "genetic scissors". One of these further developments of the CRISPR-Cas system allows genes to be activated in a targeted and efficient manner. These gene-activating CRISPR-Cas modules (CRISPRa) are based on a programmable recruitment of transcription factors to specific loci and offer several key advantages that make them particularly attractive for therapeutic applications. These advantages include inter alia low off-target effects, independence of the target gene size as well as the potential to develop gene- and mutation-independent therapeutic strategies. Herein, I will give an overview on the currently available CRISPRa modules and discuss recent developments, future potentials and limitations of this approach with a focus on therapeutic applications and in vivo delivery.

Memory efficiency in patients with drug-resistant epilepsy.

OBJECTIVE: To evaluate memory in patients with drug-resistant epilepsy. METHODS: Following an examination, 50 patients were diagnosed in accordance with the 2005 proposal of the International League Against Epilepsy and the definition of drug-resistant epilepsy from 2010. The neuropsychological examination used the Wechsler Memory Scale. It assessed seven structural types of memory: general knowledge, orientation, mental control, logical memory, number memory, associative memory, and visual reproduction. The values were compared with 50 subjects without epilepsy. RESULTS: Patients with epilepsy had statistically significantly lower values in five of seven structural units of memory. The average value of overall memory efficacy in subjects with epilepsy was 96.5 ± 19.6, while in subjects without epilepsy it was 118 ± 15.6 (p = 0.0002). Memory impairments are greater in those taking polytherapy (p = 0.0429). The overall memory efficiency correlated significantly negatively with seizure frequency (p = 0.0015) and insignificantly negative with the duration of epilepsy (p = 0.1935). CONCLUSION: Patients with drug-resistant epilepsy have lower memory efficiency. Memory impairments are greater in those taking polytherapy, as with those with more frequent seizures. The duration of epilepsy has no significant effect on overall memory performance.

THE CONCISE GUIDE TO PHARMACOLOGY 2021/22: Ion channels.

The Concise Guide to PHARMACOLOGY 2021/22 is the fifth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of nearly 1900 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes over 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/bph.15539. Ion channels are one of the six major pharmacological targets into which the Guide is divided, with the others being: G protein-coupled receptors, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2021, and supersedes data presented in the 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

Preformative, Performative and Narative in Creative Person Centered Psychopharmacotherapy.

Psychopharmacotherapy does not stand alone. The act of prescribing involves much more than solely choosing "best" medication. It seems that somewhere in the process of trying to objectify and scientify our therapy, we have neglected an important and effective dimensions of it. Psychopharmacology should consider much more than just biological dimension of drugs. Psychological, social and behavioral factors that influence drug metabolism, efficacy and side-effects are largely overlooked. Obviously, the subtext of information provided by the medical professional inevitably contains suggestion. Important part of that subtext is consisted in way we think of it, we talk of it and we perform that information. Defining of preformative and performative psychopharmacotherapy was atempted as well as desciption of narrative creative person-centered psychopharmacotherapy. Studies that indicate that medicines (SSRI) do not work on its own but as amplifier of the influence of the living conditions on mood are provided. Undirected susceptibility to change hypothesis which request acknowledging the importance of social, psychological, environmental factors to explain such the mechanisms underlying the recovery from the disease is explained. Understanding the role of medicines (SSRIs) as amplifier of the influence of the living conditions on mood represents a critical step in developing a creative, person-centered psychopharmacotherapy aimed at better matching patients with treatment and avoiding potential harmful consequences.

cAMP-dependent regulation of HCN4 controls the tonic entrainment process in sinoatrial node pacemaker cells.

It is highly debated how cyclic adenosine monophosphate-dependent regulation (CDR) of the major pacemaker channel HCN4 in the sinoatrial node (SAN) is involved in heart rate regulation by the autonomic nervous system. We addressed this question using a knockin mouse line expressing cyclic adenosine monophosphate-insensitive HCN4 channels. This mouse line displayed a complex cardiac phenotype characterized by sinus dysrhythmia, severe sinus bradycardia, sinus pauses and chronotropic incompetence. Furthermore, the absence of CDR leads to inappropriately enhanced heart rate responses of the SAN to vagal nerve activity in vivo. The mechanism underlying these symptoms can be explained by the presence of nonfiring pacemaker cells. We provide evidence that a tonic and mutual interaction process (tonic entrainment) between firing and nonfiring cells slows down the overall rhythm of the SAN. Most importantly, we show that the proportion of firing cells can be increased by CDR of HCN4 to efficiently oppose enhanced responses to vagal activity. In conclusion, we provide evidence for a novel role of CDR of HCN4 for the central pacemaker process in the sinoatrial node.

Psychological Disorders in Childhood and Adolescent Age - New Classifications.

INTRODUCTION: The eleventh revision of the International Classification of Diseases (ICD-11) is planned to be published in 2018. So called, "beta version" of the chapter of mental and behavioral disorders (ICD-11) is already available and it is considered that there will be no significant deviations in the final version. The DSM-5 was released in 2013. Changes related to mental disorders in child and adolescent psychiatry have been made in both of these classifications. To identify changes in the classifications of mental disorders in childhood and adolescent age in beta version of ICD-11 and DSM-5. METHODS: Review of mental disorders in childhood and adolescent age and their classification in ICD-11 and DSM-5. RESULTS: For disorders that are classified as "mental retardation" in ICD-10, a new term "intellectual development disorders" has been introduced in ICD-11, ie "intellectual disabilities" in DSM-5. Hyperactivity disorders and attention deficit is a separate entity in relation to ICD-10, in which it is classified as a hyperkinetic disorder. Asperger's syndrome, which is isolated from autism spectrum disorders in DSM-5, does not appear under that name in ICD-11 either. Elimination disorders are in a separate block MKB-11 and DSM-5. Speech and language disorders are classified as communication disorders in the DSM-5 classification. Selective mutism and anxiety separation disorder in childhood are in the block of anxiety and fear-related disorders in ICD-11, and among anxiety disorders in DSM-5, respectively. Reactive emotional disorder and disinhibited attachment disorder of childhood are classified as stress-related disorders in ICD-11 and DSM-5. CONCLUSIONS: The new classifications (ICD-11 and DSM-5) classify mental disorders in child and adolescent psychiatry somewhat differently from their antecedents. New entities have also been formed.

Bihavioral Addictions in Childhood and Adolescence - Pandemic Knocking Door.

INTRODUCTION: Addiction is not solely "substance dependence". Diminished control is a core defining concept of psychoactive substance addiction. Several behaviors, besides psychoactive substance ingestion, produce short-term reward that may engender diminished control over the behavior. Growing evidence suggests that behavioral addictions resemble substance addictions in many domains, including phenomenology, tolerance, comorbidity, overlapping genetic contribution, neurobiological mechanisms, and response to treatment. This similarity has given rise to the concept of non-substance or behavioral addictions, i.e., syndromes analogous to substance addiction, but with a behavioral focus. The type of excessive behaviors identified as being addictive include gambling, use of computers, playing video games, use of the internet, exercise, and shopping. Behavioral addictions have been proposed as a new class in DSM-5, but the only category included is gambling disorder. Internet gaming disorder is included in the appendix as a condition for further study. The ICD-11 included also the definition of a new disorder, gaming disorder. To present actual knowledge about behavioral addictions in childhood and adolescence. METHODS: Analysis of data in available literature in data basis and textbooks. RESULTS: Some behavioral addictions are becoming more common in children and adolescents. Dominant are gaming and gambling addiction that are also best researched. CONCLUSIONS: Behavioral addiction becomes an epidemic in children and adolescents.

A gene therapy for inherited blindness using dCas9-VPR-mediated transcriptional activation.

Catalytically inactive dCas9 fused to transcriptional activators (dCas9-VPR) enables activation of silent genes. Many disease genes have counterparts, which serve similar functions but are expressed in distinct cell types. One attractive option to compensate for the missing function of a defective gene could be to transcriptionally activate its functionally equivalent counterpart via dCas9-VPR. Key challenges of this approach include the delivery of dCas9-VPR, activation efficiency, long-term expression of the target gene, and adverse effects in vivo. Using dual adeno-associated viral vectors expressing split dCas9-VPR, we show efficient transcriptional activation and long-term expression of cone photoreceptor-specific M-opsin (Opn1mw) in a rhodopsin-deficient mouse model for retinitis pigmentosa. One year after treatment, this approach yields improved retinal function and attenuated retinal degeneration with no apparent adverse effects. Our study demonstrates that dCas9-VPR-mediated transcriptional activation of functionally equivalent genes has great potential for the treatment of genetic disorders.

Antisense Oligonucleotide- and CRISPR-Cas9-Mediated Rescue of mRNA Splicing for a Deep Intronic CLRN1 Mutation.

Mutations in CLRN1 cause Usher syndrome (USH) type III (USH3A), a disease characterized by progressive hearing impairment, retinitis pigmentosa, and vestibular dysfunction. Due to the lack of appropriate disease models, no efficient therapy for retinitis pigmentosa in USH patients exists so far. In addition, given the yet undefined functional role and expression of the different CLRN1 splice isoforms in the retina, non-causative therapies such as gene supplementation are unsuitable at this stage. In this study, we focused on the recently identified deep intronic c.254-649T>G CLRN1 splicing mutation and aimed to establish two causative treatment approaches: CRISPR-Cas9-mediated excision of the mutated intronic region and antisense oligonucleotide (AON)-mediated correction of mRNA splicing. The therapeutic potential of these approaches was validated in different cell types transiently or stably expressing CLRN1 minigenes. Both approaches led to substantial correction of the splice defect. Surprisingly, however, no synergistic effect was detected when combining both methods. Finally, the injection of naked AONs into mice expressing the mutant CLRN1 minigene in the retina also led to a significant splice rescue. We propose that both AONs and CRISPR-Cas9 are suitable strategies to initiate advanced preclinical studies for treatment of USH3A patients.

Neuropathic and cAMP-induced pain behavior is ameliorated in mice lacking CNGB1.

Cyclic nucleotide-gated (CNG) channels, which are directly activated by cAMP and cGMP, have long been known to play a key role in retinal and olfactory signal transduction. Emerging evidence indicates that CNG channels are also involved in signaling pathways important for pain processing. Here, we found that the expression of the channel subunits CNGA2, CNGA3, CNGA4 and CNGB1 in dorsal root ganglia, and of CNGA2 in the spinal cord, is transiently altered after peripheral nerve injury in mice. Specifically, we show using in situ hybridization and quantitative real-time RT-PCR that CNG channels containing the CNGB1b subunit are localized to populations of sensory neurons and predominantly excitatory interneurons in the spinal dorsal horn. In CNGB1 knockout (CNGB1-/-) mice, neuropathic pain behavior is considerably attenuated whereas inflammatory pain behavior is normal. Finally, we provide evidence to support CNGB1 as a downstream mediator of cAMP signaling in pain pathways. Altogether, our data suggest that CNGB1-positive CNG channels specifically contribute to neuropathic pain processing after peripheral nerve injury.

Enigmatic rhodopsin mutation creates an exceptionally strong splice acceptor site.

The c.620 T > G mutation in rhodopsin found in the first mapped autosomal dominant retinitis pigmentosa (adRP) locus is associated with severe, early-onset RP. Intriguingly, another mutation affecting the same nucleotide (c.620 T > A) is related to a mild, late-onset RP. Assuming that both mutations are missense mutations (Met207Arg and Met207Lys) hampering the ligand-binding pocket, previous work addressed how they might differentially impair rhodopsin function. Here, we investigated the impact of both mutations at the mRNA and protein level in HEK293 cells and in the mouse retina. We show that, in contrast to c.620 T > A, c.620 T > G is a splicing mutation, which generates an exceptionally strong splice acceptor site (SAS) resulting in a 90 bp in-frame deletion and protein mislocalization in vitro and in vivo. Moreover, we identified the core element underlying the c.620 T > G SAS strength. Finally, we demonstrate that the c.620 T > G SAS is very flexible in branch point choice, which might explain its remarkable performance. Based on these results, we suggest that (i) point mutations should be routinely tested for mRNA splicing to avoid dispensable analysis of mutations on protein level, which do not naturally exist. (ii) Puzzling disease courses of mutations in other genes might also correlate with their effects on mRNA splicing. (iii) Flexibility in branch point choice might be another factor influencing the SAS strength. (iv) The core splice element identified in this study could be useful for biotechnological applications requiring effective SAS.

Post-traumatic Stress Disorder, Metacognitions, Cognitive and Global Functioning in Bosnian War Veterans.

INTRODUCTION: Cognitive impairment is common finding in individuals with PTSD. Dysfunctional metacognitions in variety of anxiety disorders can represent generic vulnerability for anxiety disorders, as well as elements that contribute to maintaining the disorder. There is little empirical information about metacognition in war veterans with PTSD, and its relation to cognitive and/or social, occupational and psychological functioning. AIM: to determine the values and reciprocal correlations of different aspects of metacognition, with cognitive and global functioning in outpatient war veterans with PTSD. METHODS: The study was conducted on 25 war veterans (24 male), with confirmed diagnosis of PTSD by a trained psychiatrist, average age 48,5±6,2 (38-63) years, with average duration of symptoms of 9,9±4,7 (0,5-16) years. We used the Metacognitions questionnaire, Mini Mental Status Examination, and Global Assessment of Functioning Scale to assess metacognition, cognitive impairment, and global functioning. Median values of Metacognitions questionnaire subcomponents, Global Assessment of Functioning Scale and Mini Mental Status Examination were determined, and also reciprocal correlations of all parameters expressed with Spearman Rank Correlation. RESULTS: 12 patients (48%) had impaired cognitive function. Significant negative correlation of score on Mini Mental Status Examination, and negative beliefs about worry is observed (r=-0,4278, p=0,034), as well as non significant correlations between rest of metacognition subscales and score on Mini Mental Status Examination. Cognitive self-consciousness showed high positive correlation with Global Assessment of Functioning Scale (r =0,7436, p<0,0001). CONCLUSION: Follow up of metacognitions, cognitive and global functioning, and its relations, may have an important role in assessment of war veterans with posttraumatic stress disorder.

Sexual Dysfunctions in Bosnian War Veterans.

Sexual functioning of war veterans is significantly under-explored. During devastating aggression on Bosnia-Herzegovina (BiH) around 400 thousand soldiers were included in combats. It is estimated that more than 100 000 persons were killed, and more than 60 000 them were soldiers. Vast majority of them were deployed since war is ended. We found high prevalence of sexual dysfunctions in war veterans. Also significant difference in several areas of sexual functioning between war veterans with and without symptoms of posttraumatic stress disorder was found.

In Vitro Evaluation of AAV Vectors for Retinal Gene Therapy.

Gene therapy holds promise for treating previously untreatable retinal disorders. The most promising approaches use gene transfer vectors derived from adeno-associated virus (AAV) to supplement a gene function in the affected cell type. One example is gene therapy for achromatopsia which affects daylight vision. In this case, recombinant AAV (rAAV) vectors are being developed to specifically target cone photoreceptors. Development of rAAV vectors could be facilitated by the use of in vitro models. In this chapter we provide a protocol which utilizes mouse 661W cells, an in vitro model of cone photoreceptors for evaluation of the transduction efficacy of rAAV vectors.