1. Cell Communication: Hormones, Growth factors and Neurotransmitters cells can communicate with those right next to them or can communicate with targets at a distance communication can be through direct contact = adhesion-based mechanisms, transfer of materials through gap junctions or through the production of extracellular factors called signals - e.g. hormones, neurotransmitters, neuropeptides, growth factors this is called extracellular signaling these compounds exert their effects by binding to the target cells and/or entering the cell the ultimate goal is to affect the function of the cell –through modifying the expression of genes/proteins -6 steps to extracellular signaling 1.synthesis of signal (hormone, NT) 2.release of signal (exocytosis) 3.transport of signal to target (local? distance?) 4.detection of signal by target (binding to receptors) 5.change in target cell function 6.removal of the target & loss of effect -four types of extracellular chemical messangers: 1.paracrines (e.g. growth factors) 2.neurotransmitters 3.hormones 4.neurohormones (neuropeptides)

2. Extracellular Signaling: Mechanisms growth factors hormones, NTs, growth factors hormones, neuropeptides growth factors

3. Signaling within cells

4. most signals produced by cells within the body bind to receptors that are specific for that signal most receptors are found on the cell surface although some can be found within the cell! binding of the signal (ligand) to the receptor results in a series of events (signal transduction) within the cell that changes the cells function –e.g. may change the transcription rate of a gene – effects protein production Extracellular Signaling: Mechanisms

5. 1. Ion channel – binding of the S to the R changes the conformation of the R and allows transit of a specific ion -this allows entry of the signaling ion -responsible for the changing of membrane potentials -once the S is removed the “gate” closes NTs

6. 2. GPCR – binding of the signal (S) to its R activates the R -the R binds to an adjacent plasma membrane protein = adenylyl cyclase (AC) (adenylate cyclase) -AC then converts ATP to cAMP -cAMP acts as a signal within the cell = second messenger – effects cell activity -BUT some G proteins can inhibit this pathway!!!! (no AC activation, no cAMP production) HORMONES NTs

7. Growth Factors 3. Tyrosine-linked and RTKs – binding of the S to the R causes the R to dimerize (pair-up) -this activates a target (kinase) -this kinase then goes on to activate its target by phosphorylating it (adding a phosphate group) -the way the RTK and the TLK activate their targets are different but the end effect is the same d) RTKs

8. Hormones: Mechanisms of Signaling hormone producing cell = endocrine cell –e.g. thyroid, pituitary Autocrine signaling –cell responds to the hormone it produces Paracrine signaling –local action –local hormone (paracrine hormones) act on neighboring cells –autocrines act on same cell that secreted them Endocrine signaling –circulating hormones (endocrine hormones) –act on distant targets –travel in blood

9. Types of Hormones water-soluble lipid -soluble

10. Lipid-soluble Hormones Steroids –lipids derived from cholesterol –made in SER –different functional groups attached to core of structure provide uniqueness e.g. cortisol, progesterone, estrogen, testosterone, aldosterone Thyroid hormones –tyrosine ring plus attached iodines –are lipid-soluble Retinoic acid –lipids derived from retinol (vitamin A) –regulate proliferation, differentiation and death of many cell types some vitamins can acts a lipid-soluble hormones –e.g. vitamin D Nitric oxide (NO) - gas testosterone aldosterone cortisol

11. Eicosanoids –prostaglandins or leukotrienes –derived from arachidonic acid (fatty acid) –AA is converted either into prostaglandin H or into the leukotrienes –conversion of AA into prostaglandins is regulated by the COX enzymes –both act in the inflammatory reaction e.g. stimulate smooth muscle cells to contract e.g. stimulate nerve cells – pain Lipid-soluble Hormones

12. Phospholipids or Diacyl glycerol PGH3 PGH synthase -Cox-1, Cox-2 + peroxidase PGD PGE PGI Thromboxane (platelets) HPETE Leukotriene A Leukotriene C4 Leukotriene D4 Leukotriene E4 PGF -PGH synthase is comprised of a peroxidase (contains a heme group) + two Cyclooxygenase/COX isoforms – COX-1 and COX-2 -COX-1 is for the normal production of prostaglandins -COX-2 is produced by inflammatory cells -aspirin – prevents the binding of arachadonic acid to the PGH synthase catalytic site -gastric damage as a side-effect -ibuprofen – directly binds to the active site – acts a a competitor to AA -SC-558 class drugs (e.g. Vioxx) – specifically inhibit the activity of the COX-2 isoform -slow acting -longer term effects -no gastric damage

13. synthesis of steroid hormones from cholesterol backbone requires a series of specific enzymatic reactions that modifies the cholesterol –these enzymes are specific for each steroid made –they are located in specific cell types e.g. enzymes for cortisol are located specifically in the adrenal cortex not stored – once formed they released by diffusion through the PM into the blood –carrier proteins can be specific or some can pick up any steroid hormone e.g. serum albumin – indiscriminate in its steroid the hormone becomes active once released therefore the body keeps a balance of bound-inactive steroid hormones and unbound hormones that rapidly enter the cell 50% of the water soluble catecholamines are actually bound to albumin – reason is unclear only cholesterol is stored in the cytoplasm Lipid-soluble Hormones

14. Water-soluble Hormones Amine, peptide and protein hormones –modified amino acids to protein chains –serotonin, melatonin, histamine, epinephrine, insulin, dopamine –protein hormones – comprised of one or many polypeptide chains insulin, glucagon –peptide hormones – comprised of chains of amino acids e.g. growth hormone, oxytocin –amine hormones – derived from the amino acids tyrosine or tryptophan epinephrine (tyrosine and phenylalanine), serotonin (tryptophan), dopamine (tyrosine) one subcategory is called the: –catecholamines: epinephrine, norepi. and dopamine –can also act as neurotransmitters insulin

15. peptide hormones are synthesized and secreted using the same mechanism that regulates the secretion of any other protein –made as precursors in the ER – called preprohormones –transport to the Golgi where they are “pruned” to give rise to the active hormone –packaged and secreted from the Golgi –stored in the cytoplasm until needed –secretion is triggered only by specific stimulus Water-soluble Hormones

16. Action of Lipid-Soluble Hormones: Endogenous signaling Hormone must be carried by a transport protein that allows it to dissolve within the aqueous (watery) environment of the blood plasma Hormone diffuses through phospholipid bilayer & into cell the receptor is located within the cell (cytoplasm or the nucleus) binding of H to R results in its translocation into the nucleus the H then binds directly to specific sequences within the DNA = response elements this binding turns on/off specific genes – activates or inhibits gene transcription if turned on - new mRNA is formed & directs synthesis of new proteins new protein alters cell’s activity if turned off – no new protein results and the cell’s activity is altered

17. Action of Lipid-Soluble Hormones some lipid-soluble hormone don’t cross the plasma membrane – too large therefore they bind with receptors on the cell surface and trigger signaling events within the cells –signal similar to water-soluble hormones –e.g. prostaglandins

18. easily travels through the blood - hydrophilic but cannot diffuse through plasma membrane! therefore absolutely requires the expression of receptors on the cell surface – integral membrane proteins that act as first messenger the receptor protein activates a series of signaling events within the cells –e.g. epinephrine binds to receptor and activates an adjacent G-protein in membrane –G-protein activates adenylate cyclase to convert ATP to cyclic AMP (cAMP) in the cytosol –cAMP acts as a 2nd messenger –cAMP activates a series of proteins in the cytosol called kinases –kinases act to phosphorylate their targets – either activating them or inhibiting them –this speeds up/slows down physiological responses within the cell –phosphodiesterase inactivates cAMP quickly many second messengers are made in cells in response to specific hormones –e.g. calcium, IP3, DAG Cell response is turned off unless new hormone molecules arrive this mechanism allows for amplification – one H-R combination can activate two G proteins which activates 4 kinases which activate 16 more kinases etc…….

19. so the binding of a hormone to a receptor results in downstream cellular events either through direct activity of the receptor (activated by the ligand) or through production of a second messenger –types: 1. cAMP: produced by adenylyl cyclase/AC (activated by hormone G protein interaction) 2. calcium –-IP3 & DAG Action of Water-Soluble Hormones

21. cell expresses numerous type of G proteins that interact with the GPCRs –some activate adenylyl cyclase and stimulate production of cAMP – Gs (G stimulatory) –others inhibit AC – Gi (G inhibitory Gs proteinAdenylyl cyclase

22. best studied system: binding of epinephrine to the b2-adrenergic receptor activates the Gs protein and produces cAMP –Gs protein is comprised of three subunits –the active subunit is the alpha subunit –however the beta and gamma subunits have signaling roles also –note the Gs  subunit cycles between GTP and GDP bound states – called a GTPase protein –the cycling between GTP and GDP helps control its function cAMP Second Messenger systems

23. the ability to bind and hydrolyze GTP determine the function of the Gs  subunit also the site at which bacterial toxins can affect this signaling path the hydrolysis of the GTP on the Gs  protein is catalysed by the Gs  protein itself cAMP Second Messenger systems cholera stimulates the addition of an ADP onto the Gs  protein (takes it from intracellular NAD+)

24. the activity of AC is modified also by interactions with the Gi protein therefore the cell can modify its level of cAMP made by stimulating the GPCRs that activate either Gs or Gi proteins the alpha subunit of the Gi protein (Gi  ) also interacts with AC (at a different location) this Gi  protein is also an GTPase and requires the binding of GTP to become active and inhibit AC – once GTP is hydrolyzed the protein dissociates the AC inhibition is relieved –this is the basis of pertussis – the pertussis toxin prevents the hydrolysis of GTP bound to the Gi  – leads to prolonged inhibition of AC and drops in intracellular cAMP levels – inhibits cell signaling cAMP Second Messenger systems

25. synthesis of cAMP results in downstream activation of a series (i.e. a cascade) of protein kinases kinase = enzyme that phosphorylates specific amino acids (i.e. amino acid residues) on its target cAMP phosphorylates a class of kinases called cAMP-dependent protein kinases (PKAs) the cell has multiple isoforms of PKAs the PKA then phosphorylates another downstream kinase as its target these kinases can vary from cell type to cell type and also vary according to the upstream ligand –epinephrine binding catalyzes the activation of a PKA-driven cascade that catalyzes the breakdown of glycogen in liver cells – PKA activates GPK (glycogen phophorylase kinase) –insulin activated PKA which then activates acetyl CoA carboxylase and pyruvate dehydrogenase cAMP Second Messenger systems

27. Kinase cascades permit multienzyme regulation and amplify hormone signals

28. IP3 and DAG – calcium second messengers most intracellular calcium stores are sequestered in the ER or other vesicles RTK or GPCR pathways trigger the activation of phospholipase C in the PM –e.g. hormone-GPCR binding triggers activation of a Gq protein which then activates phospholipase C results in production of IP3 and DAG –IP3 diffuses through the cytoplasm and activates Ca channels within the PM or within the ER to release or allow entry of calcium within the cytoplasm –increased cytoplasmic calcium activates a class of calcium- dependent kinases called PKCs (protein kinase C) – role for DAG in this step Phosphorylation of substrates

29. Water soluble hormones and RTKs bind to protein/peptide classes of hormones –e.g. insulin –e.g. growth factors – EGF, NGF, bFGF, PDGF H binding leads to dimerization of the RTK and activation of the kinase activity endogenous to the receptor this activity phosphorylates its target and initiates the downstream signaling cascade major initiating protein is called Ras (GTPase) activation of the RTK leads to binding of the GTP-bound form of Ras this activated Ras than activates multiple downsteam paths the major one is called the MAPK pathway

30. hormones can utilize more than one receptor and more than one pathway to activate the same target –e.g. can bind and activate both GPCRs and RTKs provides the body with flexibility in its choice of hormone also allows two hormones to combine to increase the strength of an event or allows one hormone to decrease the cells response while the other hormone is trying to increase it Water-soluble Hormone Signaling: Mechanisms

31. Neurotransmitters More than 100 identified produced by neurons and stored within the neuron secrete these NTs in response to generation of an electrical signal (action potential) by the neuron bind onto target neurons (synapse) or target muscle cells (neuromuscular junction) Some bind receptors on the target and cause channels to open in the target (sodium channels) –e.g. binds to receptors on target neuron – causes generation of another action potential by target neuron –e.g. binds to receptors on target muscle cells – causes contraction Others bind receptors on the target and result in a second messenger system Results in either excitation or inhibition of the target Neuromuscular junction

32. Neurotransmitters 1. small molecules: Acetylcholine (ACh) -All neuromuscular junctions use ACh -ACh also released at chemical synapses between two neurons -Can be excitatory or inhibitory – depends on location and synapse -Inactivated by an enzyme acetylcholinesterase -Blockage of the ACh receptors by antibodies = myasthenia gravis - autoimmune disease that destroys these receptors and progressively destroys the NMJ -Anticholinesterase drugs (inhibitors of acetylcholinesterase) prevent the breakdown of ACh and raise the level that can activate the still present receptors

33. 2. Amino acids: glutamate & aspartate & GABA –most have powerful excitatory effects –Stimulate most excitatory neurons in the CNS (about ½ the neurons in the brain) –GABA (gamma amino-butyric acid) is an inhibitory neurotransmitter for 1/3 of all brain synapses i.e. inhibits the generation of an action potential by the target neuron Neurotransmitters Valium is a GABA agonist - enhancing its inhibitory effect

34. Neurotransmitters 3. Biogenic amines: modified amino acids –catecholamines: norepinephrine (NE), epinephrine, dopamine, thyroxin all derived from the amino acid tyrosine NE - role in arousal, awakening, deep sleep, regulating mood epinephrine (adrenaline) - flight or fight response dopamine - emotional responses and pleasure, decreases skeletal muscle tone –serotonin: derived from tryptophan sensory perception, temperature regulation, mood control, appetite, sleep induction feeling of well being Other types: a. ATP - released with NE from some neurons b. Nitric oxide - formed on demand in the neuron then release (brief lifespan) -role in memory and learning -produces vasodilation - Viagara enhances the effect of NO

35. Removal of Neurotransmitter Diffusion –move down concentration gradient Enzymatic degradation –acetylcholinesterase Uptake by neurons or glia cells –neurotransmitter transporters NE, epinephrine, dopamine, serotonin

36. Mimicking NTs Parkinsons - muscle stiffness due to degeneration of dopanergic nerves patients given L-Dopa (dopamine precursor) NT release can be enhanced or blocked amphetamines promote dopamine and NE release botulism causes paralysis through blockage of AcH release from motor neurons NT receptors can be blocked or activated isoproterenol binds to epinephrine receptors - used in asthma to mimic the effects of epinephrine schizophrenia - excess of dopamine Zyprexa blocks dopamine and serotonin receptors -antagonizes the effects of serotonin and dopamine NT removal can be promoted or inhibited cocaine: blocks transporters for dopamine reuptake Prozac, Paxil: blocks transporters for serotonin reuptake http://www.prozac.com/how_prozac/how_it_works.jsp?reqNavId=2.2 http://www.paxil.com/flash/depression.swf

37. Neuropeptides widespread in the nervous system excitatory and inhibitory act as hormones elsewhere in the body -Substance P -- enhances our perception of pain -opioid peptides: endorphins - release during stress, exercise enkephalins - analgesics (200x stronger than morphine) - pain-relieving effect by blocking the release of substance P dynorphins - regulates pain and emotions **acupuncture may produce loss of pain sensation because of release of opioid-like substances such as endorphins or dynorphins