M&C Pharmacology Flashcards

Question

Biosynthesis pathway of NO?

Answer 1

L-arginine -> citrulline + NO

Answer 2

Calcium enters the cell and forms complexes with calmodulin. Calcium-calmodulin complex activates MLCK which phosphorylates MLC which increases ATPase activity causing contraction. Relaxation: MLCP dephosphorylates MLC causing ATPase actvity to be reduced.

Answer 3

Acts through GPCR Ga12/13 to increase RhoA which increases ROCK which inhibits MLCP and so prevents relaxation (vasoconstrictor)

Answer 4

Shearing forces and muscarinic receptors trigger intracellular calcium increase which triggers eNOS to make NO which diffuses into smooth muscle cell immediately adjacent to endothelium and activates guanylyl cyclase to generate cGMP. cGMP interacts with PKG which opens K channels causing K to leave to cell causing hyperphosphorylation which reduces calcium channel open probability so less calcium enters the cell so less contraction takes place PKG also phosphorylates RGS2 which accelerates hydrolysis of Gq which turns down signalling by Gq coupled receptors therefore lowering calcium entry and so intracellular calcium decreases Fall in calcium inactivates MLCK PKG can also phosphorylate IRAG which is IP3 receptor associated protein causing dissociation of IRAG reducing activity of IP3 resulting in less calcium release PKG also phosphorylates MLCP which allows the cell to relax

Answer 5

Hydrolyse cGMP and/or cAMP

Answer 6

Psychogenic/reflexogenic stimuli cause neural depolarisation which causes an increase in intracellular calcium which binds to calmodulin. Calcium-calmodulin complex triggers nNOS which produces NO. NO diffuses into cells and activates soluble gunaylyl cyclase which catalyses GTP to cGMP. cGMP can now undergo 2 pathways: it can activate PKG which decreases intracellular calcium concentration which causes corpus cavernosum smooth muscle relaxation which leads to inflow of blood and therefore erection OR: PDE5 hydrolyses cGMP to 5'GMP (degrading it)

Answer 7

Binds to catalytic sites on PDE5 competing with cGMP which maintains PKG levels

Answer 8

Preferentially dilate vessels in well oxygenated areas of the lung (i.e. rematch conditions of vasodilation)

Answer 9

Promotes cell survival by inhibitinf apoptosis Turns off catalytic activity of GSK3 resulting in activation of pathways normally repressed by this enzyme (promotes proliferation, migration etc) Blocks transcription of genes normally stimulated by forkhead-related TF1

Answer 10

Degradation of PIP3 by at least 2 different phosphatases SHIP1 (restricted to meatopoietic cells) SHIP2 (ubiquitous expression) - SHIP activity removes binding sites for proteins with PIP3 selective PH domain but generates PIP2 that PKB/Akt can bind to PTEN (phosphatase) - removes both PIP3 and PIP2 which inhibits PKB/Akt

Answer 11

PH (binds PIs), EF (function unclear), X + Y (catalytic domains), C2 (calcium binding)

Answer 12

Activated RTKs bind PLC-y Non-RTKs are recruited by e.g. cytokine receptors and recruit effectors with SH2 domains e.g. PLC-y GPCRs can transactivate RTKs by e.g. Src related non-RTKs and activate PLC-y

Answer 13

1. Changes in [calcium]i (single cell populations) or functionl consequence of change 2. Accumulation of [3H]-labelled inositol phosphates against lithium block of inositol monophosphate activity - total PLC activity independent of IP3 metabolism 3. Mass of IP3 - using radioreceptor assay 4. Single cell imaging of PLC activity - biosensor for IP3/PIP2 uses a PH domain tagged with a molecule of enhanced GFP

Answer 14

``` EC = 1-2mM IC = 100nM ```

Answer 15

- Relative impermeability of the plasma membrane - Expulsion of calcium across the plasma membrane via: calcium ATPase ([Ca]i increases, Ca binds to calmodulin, Ca-calmodulin binds Ca-ATPase, Can-ATPase removes Ca) AND: NCX (Na gradient used as driving force) - Calcium buffers

Answer 16

STIM1 sits on ER membrane and has a calcium sensor (binds calcium and changes conformation based on how much is bound). When store is empty, calcium no longer binds to STIM1 allowing it to coalesce and interact with CRAC channels in the plasma membrane. This opens CRAC channels and allows calcium in right next to the ER. SERCA pumps can then pump calcium into the ER.

Answer 17

Dictylostelium discoideum - exists mainly as single cell entity moving around consuming bacteria. When it runs out of bacteria it goes into starvation mode where multiple entities congregate together to form fruiting bodies. These give off spores which land somewhere with more bacteria. Relies to some extent on GPCR signalling but has depleted complement of G protein subunits, only 1 Beta subunit - look at what happens when you remove this - still signals therefore the GPCR must signal independently of G proteins

Answer 18

cyc- lymphoma cells lack Gs protein. Desensitise protocol then reconstitute with WT lysate containg Gs proteins - still see desensitisation even without signlling pathway

Answer 19

Desensitisation correlates with phosphorylation. (e.g. by PKA or GRKs). Arrestin binds to phosphorylated receptor and causes clathrin mediated internalisation which moves receptor-arrestin complex into endosome where its fate is decided. pH in endosome more acidic than the slightly alkaline cytoplasm. The acidic environment facilitates dissociation of ligand from receptor. Also changes conformation of receptor favouring arrestin dissociation and dephosphorylation of receptor. The receptor can either be recycled to membrane or degraded - method of adjusting number of receptors available to signal

Answer 20

Conformation of arrestin (active conformation of arrestin cn determine which signalling pathway the protein scaffolds with)

Answer 21

Rhodopsin (GPCR opsin and covalently bound prosthetic group 11-cis-retinal) is present in high concentrations in the retinal rod outer segment discs. A photon of light causes isomerisation of 11-cis-retinal to all-trans-retinal, conformational change in rhodopsin and increased its GEF activity towards G protein transducin. Activated transducin interacts with PDE6 sequestering inhibitory PDEy subunits and unmasking the full catalytic activity of PDEaB. PDEaB hydrolyses cGMP leading to closure of cGMP regulated cation channels which causes decreased sodium influx - hyperpolarisation and decreased IC [calcium]

Answer 22

1. Inactivation of rhodopsin: activated rhodopsin is a substrate for rhodopsin kinase, multiple phosphorylations of the C terminal serine residues of rhodopsin occur creating high affinity binding site for rod arrestin. 2. Inactivation of transducin-GTP: RGS9-1 (GAP) 3. Re-synthesis of cGMP: guanylyl cyclase is not directly calcium sensitive but is regulated by the calcium binding protein GCAP. On illumination [calcium]i decreases, calcium dissociates from GCAP releasing it to exert its GC activating activity and increasing the rate of cGMP synthesis

Answer 23

Changes in the basal [calcium]i in the rod outer segment affects the activities GC, rhodopsin kinase and cGMP regulated cation channels through calcium sensitive regulatory proteins Changes in the basal phosphorylation of rhodopsin Translocation of key phototransduction components between ros outer and inner segments (e.g. as ambient light levels change the subcellular distribution of transducin and arrestin change rapidly and reciprocally)

Answer 24

Recoverin, GCAP and calmodulin: In its calcium bound form, recoverin binds tightly to rhodopsin kinase (GRK1) preventing it from phosphorylating rhodopsin. In its calcium bound form, GCAP is inactive. In its calcium bound form, calmodulin binds and inhibits the cGMP regulated cation channel.

Answer 25

Evidence suggests more than one C terminal site on rhodopsin must be phosphorylated by GRK1 before arrestin binds. In dark adapted rod cells, most rhodopsin is unphosphorylated, whereas with increasing levels of background illumination, rhodopsin may be in its mono or diphosphorylated form.

Answer 26

On prolonged light exposure, arrestin and RGS9-1 translocate into the outer segment while transducin and recoverin translocate out of the OS. This permits change in the ability of the phototransduction cascade to amplify and efficiently shut off signalling.

Answer 27

Allows ability to detect and report visual stimuli over a wide range of levels of background illumination

Answer 28

In cardiac myocytes, B-adrenergic receptors located in caveolae efficiently activate adenylyl cyclase 6 while prostaglandin E2 receptors (not in caveolae) do not (AC6 is solely localised in caveolae)

Answer 29

Recruited: PDGFR Stable: NMDA receptor-yotiao complex

Answer 30

SH2: binds to short peptide motifs containing pY followed by 3-6 amino acids C terminal to pY SH3: binds to proline rich peptide sequences PH: binds to PIs such as PIP2 and PIP3 PTB: binds to motifs containing pY and 3-8 amino acids N-terminal to pY PDZ: binds to last 4-5 C terminal residues in target proteins, typically ion channels and receptors You can predict protein-protein interactions

Answer 31

1. Immunosuppressive drug FK506 inactivates calcineurin by creating an unphysiological complex between calcineurin and FKBP12. Inactivation of calcineurin interrupts signals from the immune systems T cell receptor. 2. Rapamycin brings together 2 proteins that normally ignore each other (FKBP12 and FRAP). FRAP normally turns on a ribosomal kinase responsible for protein synthesis, but fails to do so when linked to rapamysin and FKBP12. This inhibits proliferation of the immune system's T lymphocytes

Answer 32

PLC A2 p90 ribosomal S6 kinase A-type K channels

Answer 33

- Ga or Gby dependent and involve conventional signalling intermediates - Some GPCRs can transactivate RTKs to act as scaffolds for Grb2/mSos recruitment and ras activation - Some nRTK such as src family kinase pyk2 and FAK are intermediates in MAPK activation by GPCRs in some cells - GPCR activation can result in proteolytic cleavage of cell surface tethered growth factor often via MMPs. The released growth factor can then act as an autocrine/paracrine activator of RTKs

Answer 34

Growth factors -> receptor -> Raf -> MKK -> ERK -> growth/differentiation Cellular stress/cytokines -> receptor -> MEKK, MLKs, ASK -> MKK -> JNK OR -> MEKK, MLK, ASK -> MKK -> p38 -> apoptosis, inflammation, differentiation

Answer 35

Wnt/B-catenin (wnt binds to LRP and frizzled causing recruitment and phosphorylation (activation) of dishevelled which disrupts the Axin/APC/CKIa/GSK-3 complex leading to GSK-3 inhibtion which prevents B-catenin from being degraded so it accumulates in the cytoplasm and translocates to the nucleus where it displaces co-repressor groucho on transcription factor TCF and recruits co-activator to cause cell proliferation) SMAD (TGFbeta binding causes TGFB receptor complex formation and activation of its type II kinase domain. Activated TGFBR type II kinase activates type I kinase domain which activates the SMAD2/3 protein which is in the cytoplasm near the membrane. SMAD4 forms a complex with 2xSMAD2/3 to form functional transcription factor. Translocates to nucleus and with the help of co-factors leads to gene transcription - causes cell growth, differentiation etc (SMAD7 blocks the formation of SMAD4-SMAD2/3 complex)) PI dependent Tubby (agonist binds to GPCR, Gq activation, activation of PLC-B, hydrolyses PIP2 -> DAG + IP3, tubby protein dislodged from membrane, nuclear translocation -> activates gene transcription)

Answer 36

``` Rounding of cell Retraction of pseudopodes Reduction of cellular and nuclear volume Nuclear fragmentation Minor modification or cytoplasmic organelles Plasma membrane blebbing Engulfment by resident phagocytes ```

Answer 37

Cytoplasmic swelling Rupture of plasma membrane Swelling of organelles Moderate chromatin condensation

Answer 38

``` (specific to skin - cells are gradually eliminated forming a dead lprotective barrier on skin surface) Elimination of cytosolic organelles Modifications to plasma membrane Accumulation of lipids in the EC space Desequamation by protease activation ```

Answer 39

Lack of chromatin condensation Massive vacuolisation of the cytoplasm Accumulation of double membraned autophagic vacuoles Little or no uptake by phagocytic cells

Answer 40

Phosphatidylserine translocates to the outside of the cell

Answer 41

Inward rectifier block by polyamines

Answer 42

Kir6.2/SUR1 - pancreatic beta cells Kir6.2/SUR2A - cardiac myocytes Kir6.2/SUR2B - smooth muscle Kir6.1/SUR2B - vascular smooth muscle

Answer 43

Katp channels are open when EC glucose is low. EC glucose increases the beta cell metabolism that increases the IC ATP levels and Katp channels close. This results in membrane depolarisation and opening of voltage gated calcium channels. Influx of calcium triggers insulin exocytosis

Answer 44

Katp channel inhibitors (e.g. glibencamide, tolbutamide) cause membrane depolarisation, calcium channel opening and insulin release

Answer 45

Tolbutamide - most specific

Answer 46

Mutations in the ATP binding site, Katp channel has reduced sensitivity to ATP so that the channel favours the open state - consequence is permanent neonatal diabetes mellitus

Answer 47

Katp channel openers e.g. diazoxide

Answer 48

Decrease hypertension by opening Katp channels in smooth muscle cells (Katp channels open causing hyperpolarisation, calcium channels close, less calcium inside the cell - relaxation) e.g. nicorandil Alopecia - Kir6.2/SUR1 and Kir6.1/SUR2B are expressed in hair follicles, Katp channel opener minoxidil activates hair growth (molecular mechanism unknown)

Answer 49

Hirsuitism - inappropriate hair growth in the normal pattern of growth Hypertrichosis - excessive hair growth outside the noraml pattern of hair growth

Answer 50

Emesis | GI function

Answer 51

Chemoreceptor trigger zone (CTZ) - not protected by blood-brain barrier allowing a chemosensory role. CTZ can be stimulated by toxins in the blood or triggered by neuronal input from the higher centres of the brain. CTZ releases 5-HT in the neuro-circuitry linking to vomiting centre to trigger the vomiting reflex GI tract - 5-HT released from enterochromaffin cellsof the intestinal mucosa stimulated by luminal stimuli e.g. food poisoning. This results in the stimulation of peripheral 5HT3 receptors on vagal afferents. This vagal afferent then stimulates local releaseof 5HT in the neuro-circuitry of the brain stem in the vomiting centre which triggers the vomiting reflex

M&C Pharmacology Flashcards

(75 cards)