133 research outputs found
Neuropeptide S-Mediated Facilitation of Synaptic Transmission Enforces Subthreshold Theta Oscillations within the Lateral Amygdala
The neuropeptide S (NPS) receptor system modulates neuronal circuit activity in
the amygdala in conjunction with fear, anxiety and the expression and extinction
of previously acquired fear memories. Using in vitro brain
slice preparations of transgenic GAD67-GFP (Δneo) mice, we investigated the
effects of NPS on neural activity in the lateral amygdala as a key region for
the formation and extinction of fear memories. We are able to demonstrate that
NPS augments excitatory glutamatergic synaptic input onto both projection
neurons and interneurons of the lateral amygdala, resulting in enhanced spike
activity of both types of cells. These effects were at least in part mediated by
presynaptic mechanisms. In turn, inhibition of projection neurons by local
interneurons was augmented by NPS, and subthreshold oscillations were
strengthened, leading to their shift into the theta frequency range. These data
suggest that the multifaceted effects of NPS on amygdaloid circuitry may shape
behavior-related network activity patterns in the amygdala and reflect the
peptide's potent activity in various forms of affective behavior and
emotional memory
Evolutionary Sequence Modeling for Discovery of Peptide Hormones
There are currently a large number of “orphan” G-protein-coupled receptors (GPCRs) whose endogenous ligands (peptide hormones) are unknown. Identification of these peptide hormones is a difficult and important problem. We describe a computational framework that models spatial structure along the genomic sequence simultaneously with the temporal evolutionary path structure across species and show how such models can be used to discover new functional molecules, in particular peptide hormones, via cross-genomic sequence comparisons. The computational framework incorporates a priori high-level knowledge of structural and evolutionary constraints into a hierarchical grammar of evolutionary probabilistic models. This computational method was used for identifying novel prohormones and the processed peptide sites by producing sequence alignments across many species at the functional-element level. Experimental results with an initial implementation of the algorithm were used to identify potential prohormones by comparing the human and non-human proteins in the Swiss-Prot database of known annotated proteins. In this proof of concept, we identified 45 out of 54 prohormones with only 44 false positives. The comparison of known and hypothetical human and mouse proteins resulted in the identification of a novel putative prohormone with at least four potential neuropeptides. Finally, in order to validate the computational methodology, we present the basic molecular biological characterization of the novel putative peptide hormone, including its identification and regional localization in the brain. This species comparison, HMM-based computational approach succeeded in identifying a previously undiscovered neuropeptide from whole genome protein sequences. This novel putative peptide hormone is found in discreet brain regions as well as other organs. The success of this approach will have a great impact on our understanding of GPCRs and associated pathways and help to identify new targets for drug development
Molecular Evolution of the Neuropeptide S Receptor
The neuropeptide S receptor (NPSR) is a recently deorphanized member of the G protein-coupled receptor (GPCR) superfamily and is activated by the neuropeptide S (NPS). NPSR and NPS are widely expressed in central nervous system and are known to have crucial roles in asthma pathogenesis, locomotor activity, wakefulness, anxiety and food intake. The NPS-NPSR system was previously thought to have first evolved in the tetrapods. Here we examine the origin and the molecular evolution of the NPSR using in-silico comparative analyses and document the molecular basis of divergence of the NPSR from its closest vertebrate paralogs. In this study, NPSR-like sequences have been identified in a hemichordate and a cephalochordate, suggesting an earlier emergence of a NPSR-like sequence in the metazoan lineage. Phylogenetic analyses revealed that the NPSR is most closely related to the invertebrate cardioacceleratory peptide receptor (CCAPR) and the group of vasopressin-like receptors. Gene structure features were congruent with the phylogenetic clustering and supported the orthology of NPSR to the invertebrate NPSR-like and CCAPR. A site-specific analysis between the vertebrate NPSR and the well studied paralogous vasopressin-like receptor subtypes revealed several putative amino acid sites that may account for the observed functional divergence between them. The data can facilitate experimental studies aiming at deciphering the common features as well as those related to ligand binding and signal transduction processes specific to the NPSR
Kinetic modeling of tricarboxylic acid cycle and glyoxylate bypass in Mycobacterium tuberculosis, and its application to assessment of drug targets
BACKGROUND: Targeting persistent tubercule bacilli has become an important challenge in the development of anti-tuberculous drugs. As the glyoxylate bypass is essential for persistent bacilli, interference with it holds the potential for designing new antibacterial drugs. We have developed kinetic models of the tricarboxylic acid cycle and glyoxylate bypass in Escherichia coli and Mycobacterium tuberculosis, and studied the effects of inhibition of various enzymes in the M. tuberculosis model. RESULTS: We used E. coli to validate the pathway-modeling protocol and showed that changes in metabolic flux can be estimated from gene expression data. The M. tuberculosis model reproduced the observation that deletion of one of the two isocitrate lyase genes has little effect on bacterial growth in macrophages, but deletion of both genes leads to the elimination of the bacilli from the lungs. It also substantiated the inhibition of isocitrate lyases by 3-nitropropionate. On the basis of our simulation studies, we propose that: (i) fractional inactivation of both isocitrate dehydrogenase 1 and isocitrate dehydrogenase 2 is required for a flux through the glyoxylate bypass in persistent mycobacteria; and (ii) increasing the amount of active isocitrate dehydrogenases can stop the flux through the glyoxylate bypass, so the kinase that inactivates isocitrate dehydrogenase 1 and/or the proposed inactivator of isocitrate dehydrogenase 2 is a potential target for drugs against persistent mycobacteria. In addition, competitive inhibition of isocitrate lyases along with a reduction in the inactivation of isocitrate dehydrogenases appears to be a feasible strategy for targeting persistent mycobacteria. CONCLUSION: We used kinetic modeling of biochemical pathways to assess various potential anti-tuberculous drug targets that interfere with the glyoxylate bypass flux, and indicated the type of inhibition needed to eliminate the pathogen. The advantage of such an approach to the assessment of drug targets is that it facilitates the study of systemic effect(s) of the modulation of the target enzyme(s) in the cellular environment
Genetic Deletion of the Nociceptin/Orphanin FQ Receptor in the Rat Confers Resilience to the Development of Drug Addiction
The nociceptin (NOP) receptor is a G-protein-coupled receptor whose natural ligand is the nociceptin/orphanin FQ (N/OFQ) peptide. Evidence from pharmacological studies suggests that the N/OFQ system is implicated in the regulation of several addiction-related phenomena, such as drug intake, withdrawal and relapse. Here, to further explore the role of NOP system in addiction, we used NOP (-/-) rats to study the motivation for cocaine, heroin and alcohol self-administration in the absence of N/OFQ function. Conditioned place preference (CPP) and saccharin (0.2% w/v) self-administration were also investigated. Results showed that NOP (-/-) rats self-administer less cocaine (0.25, 0.125 or 0.5 mg/infusion) both under a Fixed Ratio 1 and a Progressive Ratio schedule of reinforcement compared to wild type (Wt) controls. Consistently, cocaine (10 mg/kg, i.p.) was able to induce CPP in Wt but not in NOP (-/-). When NOP (-/-) rats were tested for heroin (20 μg/infusion) and ethanol (10% v/v) self-administration, they showeda significantly lower drug intake compared to Wt. Conversely, saccharin self-administration was not affected by NOP deletion, excluding the possibility of nonspecific learning deficits or generalized disruption of reward mechanisms in NOP (-/-) rats. These findings were confirmed with pharmacological experiments using two selective NOP antagonists, SB-612111 and LY2817412. Both drugs attenuated alcohol self-administration in Wt rats but not in NOP (-/-) rats. In conclusion, our results demonstrate that genetic deletion of NOP receptors confers resilience to drug abuse and support a role for NOP receptor antagonism as a potential treatment option for drug addiction.Neuropsychopharmacology accepted article preview online, 26 August 2016. doi:10.1038/npp.2016.171
The Concise Guide to PHARMACOLOGY 2023/24: G protein-coupled receptors.
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 about 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/bph.16177. G protein-coupled receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: ion channels, 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
A review on experimental and clinical genetic associations studies on fear conditioning, extinction and cognitive-behavioral treatment
Fear conditioning and extinction represent basic forms of associative learning with considerable clinical relevance and have been implicated in the pathogenesis of anxiety disorders. There is considerable inter-individual variation in the ability to acquire and extinguish conditioned fear reactions and the study of genetic variants has recently become a focus of research. In this review, we give an overview of the existing genetic association studies on human fear conditioning and extinction in healthy individuals and of related studies on cognitive-behavioral treatment (CBT) and exposure, as well as pathology development after trauma. Variation in the serotonin transporter (5HTT) and the catechol-o-methyltransferase (COMT) genes has consistently been associated with effects in pre-clinical and clinical studies. Interesting new findings, which however require further replication, have been reported for genetic variation in the dopamine transporter (DAT1) and the pituitary adenylate cyclase 1 receptor (ADCYAP1R1) genes, whereas the current picture is inconsistent for variation in the brain-derived neurotrophic factor (BDNF) gene. We end with a discussion of the findings and their limitations, as well as future directions that we hope will aid the field to develop further
Dual Role for Pilus in Adherence to Epithelial Cells and Biofilm Formation in Streptococcus agalactiae
Streptococcus agalactiae is a common human commensal and a major life-threatening pathogen in neonates. Adherence to host epithelial cells is the first critical step of the infectious process. Pili have been observed on the surface of several gram-positive bacteria including S. agalactiae. We previously characterized the pilus-encoding operon gbs1479-1474 in strain NEM316. This pilus is composed of three structural subunit proteins: Gbs1478 (PilA), Gbs1477 (PilB), and Gbs1474 (PilC), and its assembly involves two class C sortases (SrtC3 and SrtC4). PilB, the bona fide pilin, is the major component; PilA, the pilus associated adhesin, and PilC, are both accessory proteins incorporated into the pilus backbone. We first addressed the role of the housekeeping sortase A in pilus biogenesis and showed that it is essential for the covalent anchoring of the pilus fiber to the peptidoglycan. We next aimed at understanding the role of the pilus fiber in bacterial adherence and at resolving the paradox of an adhesive but dispensable pilus. Combining immunoblotting and electron microscopy analyses, we showed that the PilB fiber is essential for efficient PilA display on the surface of the capsulated strain NEM316. We then demonstrated that pilus integrity becomes critical for adherence to respiratory epithelial cells under flow-conditions mimicking an in vivo situation and revealing the limitations of the commonly used static adherence model. Interestingly, PilA exhibits a von Willebrand adhesion domain (VWA) found in many extracellular eucaryotic proteins. We show here that the VWA domain of PilA is essential for its adhesive function, demonstrating for the first time the functionality of a prokaryotic VWA homolog. Furthermore, the auto aggregative phenotype of NEM316 observed in standing liquid culture was strongly reduced in all three individual pilus mutants. S. agalactiae strain NEM316 was able to form biofilm in microtiter plate and, strikingly, the PilA and PilB mutants were strongly impaired in biofilm formation. Surprisingly, the VWA domain involved in adherence to epithelial cells was not required for biofilm formation
Multiple Polymorphisms Affect Expression and Function of the Neuropeptide S Receptor (NPSR1)
Peer reviewe
- …