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22 Cards in this Set
- Front
- Back
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T1R1+T1R3
T1R2+T1R3 T2Rs ENaC |
Umami (L-glutamine)
sugars, sweeteners (saccharin), brazzein bitter salty |
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Gustducin pathway (unconfirmed)
Chaudhari et al., 2010 |
βγ-gustducin > PLC > IP3 > Ca2+ > exocytosis of neurotransmitter
sequence similarity to transducin in rods and cones (80%). Effect on phosphodiesterases and cAMP levels |
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Taste organs and cells
Type I, II, III |
- tongue has various papillae types. no selectivity to taste at specific regions
Type I - salty taste? Type II - Taste receptor cell, sweet, unami, bitter. ATP signalling to sensory afferent fibres Type III - presynaptic cells, sour taste. 5-HT signalling to sensory afferents |
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Zhao et al., 2003
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T1r1, 2, and 3 KOs. Engineered mice expressing human T1r1 and opioid receptor in sweet taste cells
1 KO lost umami, 2KO lost sweet, 3KO lost lost. 1/3 or 2/3 KO was required for complete loss of umami/sweet taste. Human sweet preference given with human T1R1. Mice had opioid preference Both subunits required to taste sweet/umami tastes. Bitter sour and salty remained unaffected. Labelled line Taste was measured by neural response as opposed to preference/aversion, which gives good validity. Human introduction shows relevance. |
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Nie et al., 2005
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Measured interaction between Nter of T1R and and sugars
intrinsic tryptophan fluorescence (sensitive to changes in the local environment of fluorescent amino acid residues within a protein and is a good indicator of ligand binding and/or ligand-dependent conformational changes) ligand did not have fluorescence fluorescence increases in T1R2 and 3 Nter in presence of sweet ligand. but different Kd values. both bind at physiological concentrations cyclamate tastes sweet to humans, binds to hT1R3, doesn't bind with mice Now thought that most sugars bind with R2. R3 is allosterically modulated by cyclamate |
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Chandrashekar et al., 2006
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Sac locus ---> linkage analysis and genetic rescue showed that it enabled species to detect sucrose solutions v water. T1r3
T1R3 is required for both umami and sweet taste T1r2;T1r3 double knockout required to fully abolish response; response remains in single KO albeit reduced Cats are missing T1r2 gene. don't respond to sweet taste (>'.'<) |
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Zhang et al., 2003
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KO of PLCβ2 (taste specific) leads to loss of umami, sweet and bitter taste reception. salty and sour unaffected.
Rescue of PLCβ2 in bitter cells only rescues bitter taste. not sweet/umami. shows convergence of signalling, but specificity of signal by labelled line (no broad TRC tuning) |
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Xu et al., 2004
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N-terminal Venus flytrap domain of T1R2 is required for recognizing sweeteners
C-terminal transmembrane domain of T1R3 is required for recognizing sweetener cyclamate |
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O'Neill et al., 2008
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cAMP oscillation in the mouse SCN
Generating high/low cAMP levels pharmacologically eliminated circadian clock gene transcription Slowing rate of cAMP synthesis slowed rhythm Ryhthm generation. Output --> Input :O Regulate genes with CREB binding site? (Per1 and Per2) Transcirptional feedback is not always sufficient/necessary for circadian oscillation |
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O'Neill and Reddy, 2014
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Daily auto-amplification of second messenger activity, through paracrine neuropeptidergic coupling, is necessary and sufficient to account for the increased amplitude, accuracy and robustness of SCN timekeeping.
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Reverse pharmacology approach to characterising orphan GPCR
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bioinformatics and tissue expression profile
full length cloning expression in vitro puatative ligands; biological extracts applied functional screens; cAMP natural/surogate agonist, antagonist pharmaocology of receptor |
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Transcription via cAMP
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cAMP > PKA translocates to nucleus > phosphorylate CREB (cAMP response element binding protein) > CREB binds to CRE, increases transcription of target proteins
CREM and ATF1 are other proteins phosphorylated by ser/thr kinases |
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Problems with Li+
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toxic dose is near therapeutic index
Kidney tubular damage can't perfrom medicinal chemistry on it weight gain, thirst, polyuria, tremor, cognitive slowing, teratogenic (dorsal axis replication) |
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Li+ on inositol signalling
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uncompetitive inhibition of IMPase prevents InsP1 recycling back to inositol. There is a limited availability of inositol in CSF.
Inhibition of other enzymes on recycling pathway as well Berridge inositol depletion hypothesis, 1989 to account for effects on mood stabilising effects |
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Hallcher & Sherman, 1980
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Compare Lithium to other monovalent ions on IMP hydrolysis (as marker of IMPase inhibition). Only Li+ is effective, and in the therapeutic window
Crit: not actually measuring IMPase inhibition |
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Cryns, 2008
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CNS IMPase1 mouse KO
found reduced level of IMPase activity in hippocampus, frontal cortex, and cerebellum. Activity still present? other isoforms of IMPase, other enzymes capable of hydrolysing? problems with assay? |
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Glycogen synthase kinase 3 (GSK-3)
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glycogen synthesis, gene transcription, synaptic plasticity, apoptosis, cellular structure and resilience, and the circadian cycle (mood disorder implication) via Clock Genes
GSK-3β inhibits CREB and β-catenin (survival promoting growth factors) Lithium inhibits its activity |
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Gould et al., 2004
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beta-catenin is a transcription factor which is used as a marker for GSK-3beta inactivation (increased)
Lithium treatment increased beta-catenin protein levels in frontal cortex (decrease in mRNA is compensatory) GSK-3beta is phosphorylated, and so sequestered to different areas of cell. Therapeutically relevant Li+ concentrations immunochemistry to investigate? or GFP tags. beta-catenin is also structural protein - morphological changes at the synapse. |
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O'Niell et al., 2008
Cambridge |
cAMP as part of circadian clock
rhythmic and sustains the transcriptional loop of SCN (Per1::luciferase) MDL (adenyl cyclase inhibitor) dose dependent rhythm supression cAMP levels spike with PER2 expression Forskolin (adenyl cyclase activator) + IBMX (phosphodisterase inhibitor) chronic raise in [cAMP]. Causes a phase shift which can be reset by washout Epac (excchange protein activated by cAMP) inhibition causes loss of rhythm, rescued by agonist. Activates CNG? |
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Binshtok et al., 2007
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TRPV1 anatgonist + capscacin blocks Na+ current in capsaicin responsive DRG neurons
Aneasthatised to noxious thermal + mechanical stimuli, but minimal motor deficit (compared to lidocaine) Huge concentrations of antagonsit used (5mM) - get a better one? |
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Evanson et al., 2010
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GC negative feedback on PVN is mediated by endocannabinoid signalling
Dex or conjugated dex:BSA rapidly inhibited ACTH release from PVN c-Fos expression not altered. IHC shows dex stayed in PVN Dex + CB1 antagonist reverses dex-induced inhibiton of HPA axis Model: GC > membrane GC receptor > endocannab. synthesis > retrograde signal across synapse > CB1 > inhibit glutamte release. |
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Lygren et al., 2007
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Identified supramolecular complex of SERCA2, phospholamban, AKAP18-delta, and PKA on sarcoplasmic reticulum.
AKAP18-delta knockdown with siRNA interferes with phospholamban phosphorylation and adrenergic inhibition of SERCA2 AKAP organise signalosome |
