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1.
MicroPubl Biol ; 20242024.
Artigo em Inglês | MEDLINE | ID: mdl-38454952

RESUMO

Signaling by the Ral small GTPase is poorly understood in vivo . Caenorhabditis elegans animals with constitutively activated RAL-1 or deficient for the inhibitory RalGAP, HGAP-1 /2, display pale intestines. Staining with Oil Red O detected decreased intestinal lipids in the hgap-1 deletion mutant relative to the wild type. Constitutively activated RAL-1 decreased lipid detected by stimulated Raman scattering (SRS) microscopy, a label-free method of detecting lipid by laser excitation and detection. A signaling-deficient missense mutant for RAL-1 also displayed reduced lipid staining via SRS. We conclude that RAL-1 signaling regulates lipid homeostasis, biosynthesis or storage in live animals.

2.
Nat Commun ; 14(1): 8213, 2023 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-38081830

RESUMO

Achieving low-carbon development of the cement industry in the developing countries is fundamental to global emissions abatement, considering the local construction industry's rapid growth. However, there is currently a lack of systematic and accurate accounting and projection of cement emissions in developing countries, which are characterized with lower basic economic country condition. Here, we provide bottom-up quantifications of emissions from global cement production and reveal a regional shift in the main contributors to global cement CO2 emissions. The study further explores cement emissions over 2020-2050 that correspond to different housing and infrastructure conditions and emissions mitigation options for all developing countries except China. We find that cement emissions in developing countries except China will reach 1.4-3.8 Gt in 2050 (depending on different industrialization trajectories), compared to their annual emissions of 0.7 Gt in 2018. The optimal combination of low-carbon measures could contribute to reducing annual emissions by around 65% in 2050 and cumulative emissions by around 48% over 2020-2050. The efficient technological paths towards a low carbon future of cement industry vary among the countries and infrastructure scenarios. Our results are essential to understanding future emissions patterns of the cement industry in the developing countries and can inform policies in the cement sector that contribute to meeting the climate targets set out in the Paris Agreement.

3.
iScience ; 26(11): 108239, 2023 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-37953945

RESUMO

Britain has been a global leader in reducing emissions, but little progress has been made on heat, which accounts for almost one-third of UK emissions and the largest single share is domestic heat, which is responsible for 17% of the national total. Given the UK's 2050 "Net-Zero" commitment, decarbonizing heat is becoming urgent and currently one of the main pathways involves its electrification. Here, we present a spatially explicit optimization model that investigates the implications of electrifying domestic heat on the operation of the power sector. Using hourly historical gas demand data, we conclude that the domestic peak heat demand is almost 50% lower than widely cited values. A 100% electrification pathway can be achieved with only a 1.3-fold increase in generation capacity compared to a power-only decarbonization scenario, but only by leveraging the role of thermal energy storage technologies without which a further 40% increase would be needed.

4.
iScience ; 26(3): 106166, 2023 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-36994188

RESUMO

Geoengineering techniques such as solar radiation management (SRM) could be part of a future technology portfolio to limit global temperature change. However, there is public opposition to research and deployment of SRM technologies. We use 814,924 English-language tweets containing #geoengineering globally over 13 years (2009-2021) to explore public emotions, perceptions, and attitudes toward SRM using natural language processing, deep learning, and network analysis. We find that specific conspiracy theories influence public reactions toward geoengineering, especially regarding "chemtrails" (whereby airplanes allegedly spray poison or modify weather through contrails). Furthermore, conspiracies tend to spillover, shaping regional debates in the UK, USA, India, and Sweden and connecting with broader political considerations. We also find that positive emotions rise on both the global and country scales following events related to SRM governance, and negative and neutral emotions increase following SRM projects and announcements of experiments. Finally, we also find that online toxicity shapes the breadth of spillover effects, further influencing anti-SRM views.

5.
J Neurosci ; 43(14): 2597-2614, 2023 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-36898838

RESUMO

We previously demonstrated a role of piriform cortex (Pir) in relapse to fentanyl seeking after food choice-induced voluntary abstinence. Here, we used this model to further study the role of Pir and its afferent projections in fentanyl relapse. We trained male and female rats to self-administer palatable food pellets for 6 d (6 h/day) and fentanyl (2.5 µg/kg/infusion, i.v.) for 12 d (6 h/day). We assessed relapse to fentanyl seeking after 12 voluntary abstinence sessions, achieved through a discrete choice procedure between fentanyl and palatable food (20 trials/session). We determined projection-specific activation of Pir afferents during fentanyl relapse with Fos plus the retrograde tracer cholera toxin B (injected into Pir). Fentanyl relapse was associated with increased Fos expression in anterior insular cortex (AI) and prelimbic cortex (PL) neurons projecting to Pir. We next used an anatomical disconnection procedure to determine the causal role of these two projections (AI→Pir and PL→Pir) in fentanyl relapse. Contralateral but not ipsilateral disconnection of AI→Pir projections decreased fentanyl relapse but not reacquisition of fentanyl self-administration. In contrast, contralateral but not ipsilateral disconnection of PL→Pir projections modestly decreased reacquisition but not relapse. Fluorescence-activated cell sorting and quantitative PCR data showed molecular changes within Pir Fos-expressing neurons associated with fentanyl relapse. Finally, we found minimal or no sex differences in fentanyl self-administration, fentanyl versus food choice, and fentanyl relapse. Our results indicate that AI→Pir and PL→Pir projections play dissociable roles in nonreinforced relapse to fentanyl seeking versus reacquisition of fentanyl self-administration after food choice-induced voluntary abstinence.SIGNIFICANCE STATEMENT We previously showed a role of Pir in fentanyl relapse after food choice-induced voluntary abstinence in rats, a procedure mimicking human abstinence or a significant reduction in drug self-administration because of the availability of alternative nondrug rewards. Here, we aimed to further characterize the role of Pir in fentanyl relapse by investigating the role of Pir afferent projections and analyzing molecular changes in relapse-activated Pir neurons. We identified dissociable roles of two Pir afferent projections (AI→Pir and PL→Pir) in relapse to fentanyl seeking versus reacquisition of fentanyl self-administration after voluntary abstinence. We also characterized molecular changes within Pir Fos-expressing neurons associated with fentanyl relapse.


Assuntos
Fentanila , Córtex Piriforme , Humanos , Ratos , Masculino , Feminino , Animais , Ratos Sprague-Dawley , Preferências Alimentares , Alimentos , Autoadministração , Extinção Psicológica , Comportamento de Procura de Droga/fisiologia
6.
Sci Adv ; 9(2): eadd8687, 2023 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-36630511

RESUMO

High relapse rate is a key feature of opioid addiction. In humans, abstinence is often voluntary due to negative consequences of opioid seeking. To mimic this human condition, we recently introduced a rat model of incubation of oxycodone craving after electric barrier-induced voluntary abstinence. Incubation of drug craving refers to time-dependent increases in drug seeking after cessation of drug self-administration. Here, we used the activity marker Fos, muscimol-baclofen (GABAa + GABAb receptor agonists) global inactivation, Daun02-selective inactivation of putative relapse-associated neuronal ensembles, and fluorescence-activated cell sorting of Fos-positive cells and quantitative polymerase chain reaction to demonstrate a key role of vSub neuronal ensembles in incubation of oxycodone craving after voluntary abstinence, but not homecage forced abstinence. We also used a longitudinal functional magnetic resonance imaging method and showed that functional connectivity changes in vSub-related circuits predict opioid relapse after abstinence induced by adverse consequences of opioid seeking.

7.
J Neurosci ; 43(10): 1692-1713, 2023 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-36717230

RESUMO

The brain µ-opioid receptor (MOR) is critical for the analgesic, rewarding, and addictive effects of opioid drugs. However, in rat models of opioid-related behaviors, the circuit mechanisms of MOR-expressing cells are less known because of a lack of genetic tools to selectively manipulate them. We introduce a CRISPR-based Oprm1-Cre knock-in transgenic rat that provides cell type-specific genetic access to MOR-expressing cells. After performing anatomic and behavioral validation experiments, we used the Oprm1-Cre knock-in rats to study the involvement of NAc MOR-expressing cells in heroin self-administration in male and female rats. Using RNAscope, autoradiography, and FISH chain reaction (HCR-FISH), we found no differences in Oprm1 expression in NAc, dorsal striatum, and dorsal hippocampus, or MOR receptor density (except dorsal striatum) or function between Oprm1-Cre knock-in rats and wildtype littermates. HCR-FISH assay showed that iCre is highly coexpressed with Oprm1 (95%-98%). There were no genotype differences in pain responses, morphine analgesia and tolerance, heroin self-administration, and relapse-related behaviors. We used the Cre-dependent vector AAV1-EF1a-Flex-taCasp3-TEVP to lesion NAc MOR-expressing cells. We found that the lesions decreased acquisition of heroin self-administration in male Oprm1-Cre rats and had a stronger inhibitory effect on the effort to self-administer heroin in female Oprm1-Cre rats. The validation of an Oprm1-Cre knock-in rat enables new strategies for understanding the role of MOR-expressing cells in rat models of opioid addiction, pain-related behaviors, and other opioid-mediated functions. Our initial mechanistic study indicates that lesioning NAc MOR-expressing cells had different effects on heroin self-administration in male and female rats.SIGNIFICANCE STATEMENT The brain µ-opioid receptor (MOR) is critical for the analgesic, rewarding, and addictive effects of opioid drugs. However, in rat models of opioid-related behaviors, the circuit mechanisms of MOR-expressing cells are less known because of a lack of genetic tools to selectively manipulate them. We introduce a CRISPR-based Oprm1-Cre knock-in transgenic rat that provides cell type-specific genetic access to brain MOR-expressing cells. After performing anatomical and behavioral validation experiments, we used the Oprm1-Cre knock-in rats to show that lesioning NAc MOR-expressing cells had different effects on heroin self-administration in males and females. The new Oprm1-Cre rats can be used to study the role of brain MOR-expressing cells in animal models of opioid addiction, pain-related behaviors, and other opioid-mediated functions.


Assuntos
Dependência de Heroína , Heroína , Ratos , Masculino , Feminino , Animais , Heroína/farmacologia , Analgésicos Opioides/farmacologia , Núcleo Accumbens , Receptores Opioides/metabolismo , Ratos Transgênicos , Receptores Opioides mu/genética , Receptores Opioides mu/metabolismo , Dor/metabolismo
8.
MicroPubl Biol ; 20222022.
Artigo em Inglês | MEDLINE | ID: mdl-36035777

RESUMO

There exist insufficient validated "entry portal" sites in the C. elegans genome for CRISPR/Cas9-dependent insertion into endogenous genes to confer diverse spatiotemporal patterns and levels of expression on exogenous sequences. Consequently, we recognized the most common potential "entry portal" sequences: genes previously tagged with fluorescent proteins using CRISPR/Cas9. As proof of concept, we used existing mKate2-encoding sequences inserted in the 5' end of genes as an insertion point for the auxin inducible degron, AID*. This sequence permits reasonably efficient insertion that can be employed using a variety of approaches for different end goals. Our strategy is thus generalizable to many needs.

9.
eNeuro ; 9(4)2022.
Artigo em Inglês | MEDLINE | ID: mdl-35768212

RESUMO

The orbitofrontal cortex (OFC) and piriform cortex (Pir) play a role in fentanyl relapse after food choice-induced voluntary abstinence, a procedure mimicking abstinence because of availability of alternative nondrug rewards. We used in situ hybridization and pharmacology to determine the role of OFC and Pir cannabinoid and dopamine receptors in fentanyl relapse. We trained male and female rats to self-administer food pellets for 6 d (6 h/d) and intravenous fentanyl (2.5 µg/kg/infusion) for 12 d (6 h/d). We assessed fentanyl relapse after 12 discrete choice sessions between fentanyl and food (20 trials/d), in which rats voluntarily reduced fentanyl self-administration. We used RNAscope to determine whether fentanyl relapse is associated with activity (indicated by Fos) in OFC and Pir cells expressing Cnr1 [which encodes cannabinoid 1 (CB1) receptors] or Drd1 and Drd2 (which encode dopamine D1 and D2 receptors). We injected a CB1 receptor antagonist or agonist (0.3 or 1.0 µg AM251 or WIN55,212-2/hemisphere) into OFC or a dopamine D1 receptor antagonist (1.0 or 3.0 µg SCH39166/hemisphere) into Pir to determine the effect on fentanyl relapse. Fentanyl relapse was associated with OFC cells co-expressing Fos and Cnr1 and Pir cells co-expressing Fos and Drd1 However, injections of the CB1 receptor antagonist AM251 or agonist WIN55,212-2 into OFC or the dopamine D1 receptor antagonist SCH39166 into Pir had no effect on fentanyl relapse. Fentanyl relapse is associated with activation of Cnr1-expressing OFC cells and Drd1-expressing Pir cells, but pharmacological manipulations do not support causal roles of OFC CB1 receptors or Pir dopamine D1 receptors in fentanyl relapse.


Assuntos
Canabinoides , Córtex Piriforme , Animais , Canabinoides/farmacologia , Dopamina , Antagonistas de Dopamina/farmacologia , Feminino , Fentanila/farmacologia , Masculino , Ratos , Receptor CB1 de Canabinoide , Receptores de Dopamina D1/metabolismo , Recidiva
10.
G3 (Bethesda) ; 12(6)2022 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-35551383

RESUMO

Development of the Caenorhabditis elegans vulva is a classic model of organogenesis. This system, which starts with 6 equipotent cells, encompasses diverse types of developmental event, including developmental competence, multiple signaling events to control precise and faithful patterning of three cell fates, execution and proliferation of specific cell lineages, and a series of sophisticated morphogenetic events. Early events have been subjected to extensive mutational and genetic investigations and later events to cell biological analyses. We infer the existence of dramatically changing profiles of gene expression that accompanies the observed changes in development. Yet, except from serendipitous discovery of several transcription factors expressed in dynamic patterns in vulval lineages, our knowledge of the transcriptomic landscape during vulval development is minimal. This study describes the composition of a vulva-specific transcriptome. We used tissue-specific harvesting of mRNAs via immunoprecipitation of epitope-tagged poly(A) binding protein, PAB-1, heterologously expressed by a promoter known to express GFP in vulval cells throughout their development. The identified transcriptome was small but tightly interconnected. From this data set, we identified several genes with identified functions in development of the vulva and validated more with promoter-GFP reporters of expression. For one target, lag-1, promoter-GFP expression was limited but a fluorescent tag of the endogenous protein revealed extensive expression. Thus, we have identified a transcriptome of C. elegans vulval lineages as a launching pad for exploration of functions of these genes in organogenesis.


Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Ligação a DNA/genética , Feminino , Morfogênese , Transcriptoma , Vulva/metabolismo
11.
Methods Mol Biol ; 2438: 345-376, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35147953

RESUMO

The Caenorhabditis elegans embryo is well suited for analysis of directed cell rearrangement via modern microscopy, due to its simple organization, short generation time, transparency, invariant lineage, and the ability to generate engineered embryos expressing various fluorescent proteins. This chapter provides an overview of routine microscopy techniques for imaging dorsal intercalation, a convergent extension-like morphogenetic movement in the embryonic epidermis of C. elegans, including making agar mounts, low-cost four-dimensional (4D) Nomarski microscopy, laser microsurgery, and 4D fluorescence microscopy using actin and junctional fusion proteins, as well as tissue-specific promoters useful for studying dorsal intercalation.


Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Embrião não Mamífero/metabolismo , Células Epidérmicas/metabolismo , Epiderme/metabolismo , Microscopia de Fluorescência , Morfogênese
12.
Small GTPases ; 13(1): 128-135, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-33956571

RESUMO

Ras is the most mutated oncoprotein in cancer. Among the three oncogenic effectors of Ras - Raf, PI3 Kinase and RalGEF>Ral - signalling through RalGEF>Ral (Ras-like) is by far the least well understood. A variety of signals and binding partners have been defined for Ral, yet we know little of how Ral functions in vivo. This review focuses on previous research in Drosophila that defined a function for Ral in apoptosis and established indirect relationships among Ral, the CNH-domain MAP4 Kinase misshapen, and the JNK MAP kinase basket. Most of the described signalling components are not essential in C. elegans, facilitating subsequent analysis using developmental patterning of the C. elegans vulval precursor cells (VPCs). The functions of two paralogous CNH-domain MAP4 Kinases were defined relative to Ras>Raf, Notch and Ras>RalGEF>Ral signalling in VPCs. MIG-15, the nematode ortholog of misshapen, antagonizes both the Ral-dependent and Ras>Raf-dependent developmental outcomes. In contrast, paralogous GCK-2, the C. elegans ortholog of Drosophila happyhour, propagates the 2°-promoting signal of Ral. Manipulations via CRISPR of Ral signalling through GCK-2 coupled with genetic epistasis delineated a Ras>RalGEF>Ral>Exo84>GCK-2>MAP3KMLK-1> p38PMK-1 cascade. Thus, genetic analysis using invertebrate experimental organisms defined a cascade from Ras to p38 MAP kinase.


Assuntos
Caenorhabditis elegans , Transdução de Sinais , Animais , Caenorhabditis elegans/metabolismo , Transdução de Sinais/genética , Fosfatidilinositol 3-Quinases/metabolismo , Drosophila/metabolismo
14.
G3 (Bethesda) ; 11(9)2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34544135

RESUMO

Using model organisms to identify novel therapeutic targets is frequently constrained by pre-existing genetic toolkits. To expedite positive selection for identification of novel downstream effectors, we engineered conditional expression of activated CED-10/Rac to disrupt Caenorhabditis elegans embryonic morphogenesis, titrated to 100% lethality. The strategy of engineering thresholds for positive selection using experimental animals was validated with pharmacological and genetic suppression and is generalizable to diverse molecular processes and experimental systems.


Assuntos
Proteínas de Caenorhabditis elegans , Animais , Caenorhabditis elegans/genética
15.
J Cell Sci ; 134(17)2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34341823

RESUMO

The extracellular signal-regulated kinases (ERKs) are mitogen-activated protein kinases (MAPKs) that are utilized downstream of Ras to Raf to MEK signaling to control activation of a wide array of targets. Activation of ERKs is elevated in Ras-driven tumors and RASopathies, and thus is a target for pharmacological inhibition. Regulatory mechanisms of ERK activation have been studied extensively in vitro and in cultured cells, but little in living animals. In this study, we tagged the Caenorhabditis elegans ERK-encoding gene, mpk-1. MPK-1 is ubiquitously expressed with elevated expression in certain contexts. We detected cytosol-to-nuclear translocation of MPK-1 in maturing oocytes and hence validated nuclear translocation as a reporter of some activation events. During patterning of vulval precursor cells (VPCs), MPK-1 is necessary and sufficient for the central cell, P6.p, to assume the primary fate. Yet MPK-1 translocates to the nuclei of all six VPCs in a temporal and concentration gradient centered on P6.p. This observation contrasts with previous results using the ERK nuclear kinase translocation reporter of substrate activation, raising questions about mechanisms and indicators of MPK-1 activation. This system and reagent promise to provide critical insights into the regulation of MPK-1 activation within a complex intercellular signaling network.


Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Feminino , Proteína Quinase 1 Ativada por Mitógeno , Fosfotransferases , Transdução de Sinais , Vulva
16.
Pharmacol Rev ; 73(3): 1050-1083, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34257149

RESUMO

Relapse to drug use during abstinence is a defining feature of addiction. During the last several decades, this clinical scenario has been studied at the preclinical level using classic relapse/reinstatement models in which drug seeking is assessed after experimenter-imposed home-cage forced abstinence or extinction of the drug-reinforced responding in the self-administration chambers. To date, however, results from studies using rat relapse/reinstatement models have yet to result in Food and Drug Administration-approved medications for relapse prevention. The reasons for this state of affairs are complex and multifaceted, but one potential reason is that, in humans, abstinence is often self-imposed or voluntary and occurs either because the negative consequences of drug use outweigh the drug's rewarding effects or because of the availability of nondrug alternative rewards that are chosen over the drug. Based on these considerations, we and others have recently developed rat models of relapse after voluntary abstinence, achieved either by introducing adverse consequences to drug taking (punishment) or seeking (electric barrier) or by providing mutually exclusive choices between the self-administered drug and nondrug rewards (palatable food or social interaction). In this review, we provide an overview of these translationally relevant relapse models and discuss recent neuropharmacological findings from studies using these models. We also discuss sex as a biological variable, future directions, and clinical implications of results from relapse studies using voluntary abstinence models. Our main conclusion is that the neuropharmacological mechanisms controlling relapse to drug seeking after voluntary abstinence are often different from the mechanisms controlling relapse after home-cage forced abstinence or reinstatement after extinction. SIGNIFICANCE STATEMENT: This review describes recently developed rat models of relapse after voluntary abstinence, achieved either by introducing adverse consequences to drug taking or seeking or by providing mutually exclusive choices between the self-administered drug and nondrug rewards. This review discusses recent neuropharmacological findings from studies using these models and discusses future directions and clinical implications.


Assuntos
Fissura , Preparações Farmacêuticas , Animais , Comportamento de Procura de Droga , Humanos , Modelos Animais , Ratos , Recidiva , Autoadministração
17.
MicroPubl Biol ; 20212021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34142023

RESUMO

The C. elegans dauer is an alternative third stage larva induced by dense population and adverse environmental conditions. Genes whose mutants caused dauer formation constitutive (Daf-c) and dauer formation defective (Daf-d) phenotypes were ordered via epistasis into a signaling network, with upstream DAF-7/TGF-beta and DAF-11/receptor guanylyl cyclase defining sensory branches and downstream DAF-2/Insulin receptor and DAF-12/nuclear hormone receptor executing the dauer decision. Mutations in the Scd genes were defined as incompletely penetrant suppressors of the constitutive dauer phenotype conferred by mutation of the DAF-7/TGF-beta signaling axis. SCD-2 was previously shown to be an ortholog of mammalian ALK (Anaplastic Lymphoma Kinase), a receptor tyrosine kinase. Mutations disrupting the HEN-1/Jeb ligand, SOC-1/DOS/GAB adaptor protein and SMA-5/ERK5 atypical MAP Kinase caused Scd phenotypes similar to that of mutant SCD-2. This group regulated expression from a TGF-beta-responsive GFP reporter. Here we find that a strain harboring a mutation in the uncharacterized SCD-4 is mutant for MLK-1, the C. elegans ortholog of mammalian Mixed Lineage Kinase and Drosophila slipper (slpr), a MAP3 kinase. We validated this finding by showing that a previously characterized deletion in MLK-1 caused a Scd phenotype similar to that of mutant SCD-4 and altered expression from the TGF-beta-responsive GFP reporter, suggesting that SCD-4 and MLK-1 are the same protein. Based on shared phenotypes and molecular identities, we hypothesize that MLK-1 functions as a MAP3K in the SCD-2/ALK cascade that signals through SMA-5/ERK5 MAP Kinase to modulate the output of the TGF-beta cascade controlling dauer formation in response to environmental cues.

18.
Methods Mol Biol ; 2262: 423-436, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33977493

RESUMO

Characterizing the consequences of mutated Ras/LET-60 on the development of the C. elegans vulva has provided critical insights into the role of Ras in normal animal development. Furthermore, double mutant analysis revealed the role of Ras relative to other components of growth factor signal transduction. Here we describe the combined use of principles of parallelism and epistasis to investigate the use of different Ras effectors, Raf and RalGEF > Ral, during the development of the vulva and other tissues. We additionally describe the use of these principles to delineate the function of the close Ras relative, RAP-1. The worm continues to lead the way in clarifying otherwise poorly understood functions of Ras during animal development.


Assuntos
Caenorhabditis elegans/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Vulva/crescimento & desenvolvimento , Proteínas ral de Ligação ao GTP/metabolismo , Proteínas rap1 de Ligação ao GTP/metabolismo , Proteínas ras/metabolismo , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Feminino , Transdução de Sinais , Vulva/metabolismo , Proteínas ral de Ligação ao GTP/genética , Proteínas rap1 de Ligação ao GTP/genética , Proteínas ras/genética
19.
Dev Biol ; 477: 37-48, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33991533

RESUMO

Ras is the most commonly mutated oncogene in humans and uses three oncogenic effectors: Raf, PI3K, and RalGEF activation of Ral. Understanding the importance of RalGEF>Ral signaling in cancer is hampered by the paucity of knowledge about their function in animal development, particularly in cell movements. We found that mutations that disrupt function of RalGEF or Ral enhance migration phenotypes of mutants for genes with established roles in cell migration. We used as a model the migration of the canal associated neurons (CANs), and validated our results in HSN cell migration, neurite guidance, and general animal locomotion. These functions of RalGEF and Ral are specific to their control of Ral signaling output rather than other published functions of these proteins. In this capacity Ral functions cell autonomously as a permissive developmental signal. In contrast, we observed Ras, the canonical activator of RalGEF>Ral signaling in cancer, to function as an instructive signal. Furthermore, we unexpectedly identified a function for the close Ras relative, Rap1, consistent with activation of RalGEF>Ral. These studies define functions of RalGEF>Ral, Rap1 and Ras signaling in morphogenetic processes that fashion the nervous system. We have also defined a model for studying how small GTPases partner with downstream effectors. Taken together, this analysis defines novel molecules and relationships in signaling networks that control cell movements during development of the nervous system.


Assuntos
Proteínas de Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/fisiologia , Fatores de Troca do Nucleotídeo Guanina/fisiologia , Sistema Nervoso/fisiopatologia , Transdução de Sinais , Proteínas ral de Ligação ao GTP/fisiologia , Proteínas ras/fisiologia , Animais , Sistemas CRISPR-Cas , Caenorhabditis elegans/embriologia , Indução Embrionária , Genes ras , Sistema Nervoso/embriologia , Neurônios/fisiologia , Proteínas ras/genética
20.
Psychopharmacology (Berl) ; 238(7): 1885-1897, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33765177

RESUMO

RATIONALE AND OBJECTIVE: Pain-related factors increase the risk for opioid addiction, and pain may function as a negative reinforcer to increase opioid taking and seeking. However, experimental pain-related manipulations generally do not increase opioid self-administration in rodents. This discrepancy may reflect insufficient learning of pain-relief contingencies or confounding effects of pain-related behavioral impairments. Here, we determined if pairing noxious stimuli with opioid self-administration would promote pain-related reinstatement of opioid seeking or increase opioid choice over food. METHODS: In Experiment 1, rats self-administered fentanyl in the presence or absence of repeated intraplantar capsaicin injections in distinct contexts to model context-specific exposure to cutaneous nociception. After capsaicin-free extinction in both contexts, we tested if capsaicin would reinstate fentanyl seeking. In Experiment 2, rats self-administered heroin after intraperitoneal (i.p.) lactic acid injections to model acute visceral inflammatory pain. After lactic acid-free extinction, we tested if lactic acid would reinstate heroin seeking. In Experiment 3, we tested if repeated i.p. lactic acid or intraplantar Complete Freund's Adjuvant (CFA; to model sustained inflammatory pain) would increase fentanyl choice over food. RESULTS: In Experiments 1-2, neither capsaicin nor lactic acid reinstated opioid seeking after extinction, and lactic acid did not increase heroin-induced reinstatement. In Experiment 3, lactic acid and CFA decreased reinforcement rate without affecting fentanyl choice. CONCLUSIONS: Results extend the range of conditions across which pain-related manipulations fail to increase opioid seeking in rats and suggest that enhanced opioid-addiction risk in humans with chronic pain involves factors other than enhanced opioid reinforcement and relapse.


Assuntos
Analgésicos Opioides/administração & dosagem , Comportamento de Escolha/efeitos dos fármacos , Comportamento de Procura de Droga/efeitos dos fármacos , Medição da Dor/psicologia , Dor/psicologia , Reforço Psicológico , Animais , Comportamento de Escolha/fisiologia , Condicionamento Operante/efeitos dos fármacos , Condicionamento Operante/fisiologia , Comportamento de Procura de Droga/fisiologia , Extinção Psicológica/efeitos dos fármacos , Extinção Psicológica/fisiologia , Feminino , Fentanila/farmacologia , Masculino , Transtornos Relacionados ao Uso de Opioides/psicologia , Dor/tratamento farmacológico , Medição da Dor/métodos , Ratos , Autoadministração/métodos
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