cell biology 7 Flashcards
G protein–coupled receptors (GPCRs)
a large protein family of receptors that sense molecules outside the cell and activate inside signal transduction pathways and, ultimately, cellular responses. They are called seven-transmembrane receptors because they pass through the cell membrane seven times. The ligands that bind and activate these receptors include light-sensitive compounds, odors, pheromones, hormones, and neurotransmitters, and vary in size from small molecules to peptides to large proteins. G protein–coupled receptors are involved in many diseases, and are also the target of approximately 40% of all modern medicinal drugs. There are two principal signal transduction pathways involving the G protein–coupled receptors: the cAMP signal pathway and the phosphatidylinositol signal pathway. When a ligand binds to the GPCR it causes a conformational change in the GPCR, which allows it to act as a guanine nucleotide exchange factor (GEF). The GPCR can then activate an associated G protein by exchanging its bound GDP for a GTP. The G protein’s α subunit, together with the bound GTP, can then dissociate from the β and γ subunits to further affect intracellular signaling proteins or target functional proteins directly depending on the α subunit type. GPCR tend to dimerize but it is not clear if this is necessary to function.
GPCRs structure
GPCRs are integral membrane proteins that possess seven membrane-spanning domains or transmembrane helices. The extracellular parts of the receptor can be glycosylated. These extracellular loops also contain two highly conserved cysteine residues that form disulfide bonds to stabilize the receptor structure. Some seven-transmembrane helix proteins (channelrhodopsin) that resemble GPCRs may contain ion channels, within their protein.
GPCRs structure-function relationship
In terms of structure, GPCRs are characterized by an extracellular N-terminus, followed by seven transmembrane (7-TM) α-helices (TM-1 to TM-7) connected by three intracellular (IL-1 to IL-3) and three extracellular loops (EL-1 to EL-3), and finally an intracellular C-terminus. The GPCR arranges itself into a tertiary structure resembling a barrel, with the seven transmembrane helices forming a cavity within the plasma membrane that serves a ligand-binding domain that is often covered by EL-2. Ligands may also bind elsewhere, however, as is the case for bulkier ligands (e.g., proteins or large peptides), which instead interact with the extracellular loops, or, as illustrated by the class C metabotropic glutamate receptors (mGluRs), the N-terminal tail. The class C GPCRs are distinguished by their large N-terminal tail, which also contains a ligand-binding domain. Upon glutamate-binding to an mGluR, the N-terminal tail undergoes a conformational change that leads to its interaction with the residues of the extracellular loops and TM domains. The eventual effect of all three types of agonist-induced activation is a change in the relative orientations of the TM helices (likened to a twisting motion) leading to a wider intracellular surface and “revelation” of residues of the intracellular helices and TM domains crucial to signal transduction function (i.e., G-protein coupling). Inverse agonists and antagonists may also bind to a number of different sites, but the eventual effect must be prevention of this TM helix reorientation. The structure of the N- and C-terminal tails of GPCRs may also serve important functions beyond ligand-binding. For example, The C-terminus of M3 muscarinic receptors is sufficient, and the six-amino-acid polybasic (KKKRRK) domain in the C-terminus is necessary for its preassembly with Gq proteins.[35] In particular, the C-terminus often contains serine (Ser) or threonine (Thr) residues that, when phosphorylated, increase the affinity of the intracellular surface for the binding of scaffolding proteins called β-arrestins (β-arr).[36] Once bound, β-arrestins both sterically prevent G-protein coupling and may recruit other proteins, leading to the creation of signaling complexes involved in extracellular-signal regulated kinase (ERK) pathway activation or receptor endocytosis (internalization). As the phosphorylation of these Ser and Thr residues often occurs as a result of GPCR activation, the β-arr-mediated G-protein-decoupling and internalization of GPCRs are important mechanisms of desensitization.
GPCRs ligand binding
the ligands of GPCRs typically bind within the transmembrane domain. However, protease-activated receptors are activated by cleavage of part of their extracellular domain.
GPCRs conformation change
It is known that in the inactive state, the GPCR is bound to a heterotrimeric G protein complex. Binding of an agonist to the GPCR results in a conformation change in the receptor that is transmitted to the bound Gα subunit of the heterotrimeric G protein. The activated Gα subunit exchanges GTP in place of GDP which in turn triggers the dissociation of Gα subunit from the Gβγ dimer and from the receptor. The dissociated Gα and Gβγ subunits interact with other intracellular proteins to continue the signal transduction cascade while the freed GPCR is able to rebind to another heterotrimeric G protein to form a new complex that is ready to initiate another round of signal transduction.
hetertimeric G protein
These proteins are activated by G protein-coupled receptors and are made up of alpha (α), beta (β) and gamma (γ) subunits, the latter two referred to as the beta-gamma complex. Gα subunits consist of two domains, the GTPase domain (the deactivating part), and the alpha-helical domain. The β and γ subunits are closely bound to one another and are referred to as the beta-gamma complex. Upon activation of the GPCR, the Gβγ complex is released from the Gα subunit after its GDP-GTP exchange. The free Gβγ? complex can act as a signaling molecule itself, by activating other second messengers or by gating ion channels directly. The GDP dissociation is the rate limiting step and the conformation change in GPCR enzymaticaly favors this dissociation. The high concentration of GTP in the cytoplasm ensures that GTP will replace GDP.
G-protein activation/deactivation cycle
When the receptor is inactive, the GEF domain may be bound to an also inactive α-subunit of a heterotrimeric G-protein. These “G-proteins” are a trimer of α, β, and γ subunits (known as Gα, Gβ, and Gγ, respectively) that is rendered inactive when reversibly bound to Guanosine diphosphate (GDP) (or, alternatively, no guanine nucleotide) but active when bound to Guanosine triphosphate (GTP). Upon receptor activation, the GEF domain, in turn, allosterically activates the G-protein by facilitating the exchange of a molecule of GDP for GTP at the G-protein’s α-subunit. The cell maintains a 10:1 ratio of cytosolic GTP:GDP so exchange for GTP is ensured. At this point, the subunits of the G-protein dissociate from the receptor, as well as each other, to yield a Gα-GTP monomer and a tightly interacting Gβγ dimer, which are now free to modulate the activity of other intracellular proteins. The extent to which they may diffuse, however, is limited due to the palmitoylation of Gα and the presence of an isoprenoid moiety that has been covalently added to the C-termini of Gγ. Because Gα also has slow GTP→GDP hydrolysis capability, the inactive form of the α-subunit (Gα-GDP) is eventually regenerated, thus allowing reassociation with a Gβγ dimer to form the “resting” G-protein, which can again bind to a GPCR and await activation. The rate of GTP hydrolysis is often accelerated due to the actions of another family of allosteric modulating proteins called Regulators of G-protein Signaling, or RGS proteins, which are a type of GTPase-Activating Protein, or GAP. In fact, many of the primary effector proteins (e.g., adenylate cyclases) that become activated/inactivated upon interaction with Gα-GTP also have GAP activity. Thus, even at this early stage in the process, GPCR-initiated signaling has the capacity for self-termination.
Guanine nucleotide exchange factors (GEFs)
activate monomeric GTPases by stimulating the release of guanosine diphosphate (GDP) to allow binding of guanosine triphosphate (GTP).
Kinase
What they really do is adding phosphate groups (bulky, negatively charged) to proteins, which in turn can regulate their activity. The action of kinases (phosphorylation) is opposed by the action of phosphatases (dephosphorylation). Kinases can be classified based on the amino acid residue they phosphorylate (Ser and Thr; or Tyr), their substrate, their activation, or their phylogenetic relationship. There are much more kinases (388 S/T; 518 overall) than phosphatases (38 S/T; ~130 overall), meaning that a single phosphatase has to oppose the action of many kinases. This has lead to the traditional view that phosphatases are less specific and less regulated; this may be true in comparison to many kinases, but there is both substrate selectivity and regulation of activity among phosphatases. Ser/Thr and Tyr have phosphorylatable hydroxyl (-OH) groups, and chemically, a phosphorylation reaction catalyzed by a kinase is a nucleophilic attack of such hydroxyl group onto the γ-phosphate of an ATP molecule. A kinase catalyzes this by promoting an ideal positioning of the reaction partners.
Structure and Regulation of Protein Kinases
A kinase domain consists of a small and large. ATP binds in the cleft between the lobes; interaction of the substrate is usually mostly with the large lobe. A “closed conformation” of the glycine rich loop in the small lobe forces the γ-Phosphate of the ATP into the right position for phosphorylation (a fast reaction). An “open conformation” of the glycine rich loop then allow exchange of the generated ADP for a new ATP (a slow reaction). Thus, kinase activity is thought to require alternating open and closed conformations. The active conformation of all kinases is highly conserved. This presents a problem for making specific inhibitors for individual kinases. However, the inactive conformations are not, since there are many ways to distort conformation to prevent activity (an opportunity for specific inhibitors, now also realized by the pharmaceutical industry). Generally, the ATP binding pocket is somehow distorted in inactive conformations (glycine rich loop; C-helix; activation loop). In many but not all kinases, the activation loop has to be phosphorylated for full activity. Another common regulatory theme is block of the active site by an inhibitory “pseudo-substrate” sequence.
MAP kinase cascades
are examples how kinases are activated by other “upstream” kinases. MAP kinase stands for “mitogen activated protein kinase”, but the activating “mitogen” (something that induces cell division) is far upstream in the activation… and this upstream signal does not even have to be a mitogen.
Calcineurin
a protein phosphatase also known as protein phosphatase 3, and calcium-dependent serine-threonine phosphatase. also called Protein Phosphatase 2B. When an antigen-presenting cell interacts with a T cell receptor on T cells, there is an increase in the cytoplasmic level of calcium, which activates calcineurin, by binding a regulatory subunit and activating calmodulin binding. Calcineurin induces different transcription factors (NFATs) that are important in the transcription of IL-2 genes. IL-2 activates T-helper lymphocytes and induces the production of other cytokines. In this way, it governs the action of cytotoxic lymphocytes. The amount of IL-2 being produced by the T-helper cells is believed to influence the extent of the immune response significantly. Its activity is inhibited by both cyclosporin and tacrolimus. Both bind to an “immunophilin” (cyclophilin for cylcosporin; FKBP-12 = FK506 binding protein for tacrolimus) which then inhibits phosphatase activity of Calcineurin.
mTOR
Sirolimus = rapamycin = rapamune: immunosuppressant; can be used in combination with additional drugs in case of high risk for calcineurin inhibitor associated nephrotoxicity. Binds to FKBP-12 which then inhibits kinase activity of mTOR (not phosphatase activity of calcineurin)
cyclosporin
immunosuppressant indicated against graft rejection after transplantation. Ciclosporin binds to the cytosolic protein cyclophilin (immunophilin) of lymphocytes, especially T cells. This complex of ciclosporin and cyclophilin inhibits calcineurin, which, under normal circumstances, is responsible for activating the transcription of interleukin 2.
rapamycin
Unlike the similarly named tacrolimus, sirolimus is not a calcineurin inhibitor, but it has a similar suppressive effect on the immune system. Sirolimus inhibits the the production of interleukin-2 (IL-2) via action on mTOR, and thereby blocks activation of T and B cells. Binds to FKBP-12 which then inhibits kinase activity of mTOR (not phosphatase activity of calcineurin)
Kinases as pharmacology targets
Calcineurin (=Protein Phosphatase 2B) and mTOR (a Ser/Thr kinase) inhibitors (cyclosporin and rapamycin) are important immuno-suppressants. A Tyr kinase is the target of Gleevac, a highly successful drug in therapy of chronic myeloid leukemia.
CaMKII
a serine/threonine-specific protein kinase that is regulated by the Ca2+/calmodulin complex. The sensitivity of the CaMKII enzyme to calcium and calmodulin is governed by the variable and self-associative domains. This sensitivity level of CaMKII will also modulate the different states of activation for the enzyme. Initially, the enzyme is activated; however, autophosphorylation does not occur because there is not enough Calcium or calmodulin present to bind to neighboring subunits. As greater amounts of calcium and calmodulin accumulate, autophosphorylation occurs leading to persistent activation of the CaMKII enzyme for a short period of time. Low frequency stimulation (low Ca) calcineurin wins, it is more sensitive, leading to long term depression. High frequency stimulation -> high Ca2+ -> CaMKII wins (gets more sensitive in the process).
regulators of synaptic strength
CaMKII and Calcineurin (a Ca2+ regulated kinase and phosphate, respectively) are important mediators of long term potentiation and long term depression (LTP and LTD) of synaptic strength, respectively. (These forms of changes in the strength of connection between neurons are thought to underlie higher brain functions, especially learning and memory). (Slides 17+18). This is another example of Ca2+ as a node in signal transduction; depending on the pathway it activates, it can have opposite cellular effects.
Calcium signaling
Calcium can act in signal transduction resulting from activation of ion channels or as a second messenger caused by indirect signal transduction pathways such as G protein-coupled receptors.
Calcium signaling through ion channels
Movement of calcium ions from the extracellular compartment to the intracellular compartment alters membrane potential. This is seen in the heart, during the plateau phase of ventricular contraction. In this example, calcium acts to maintain depolarization of the heart. Calcium signaling through ion channels is also important in neuronal synaptic transmission.
Calcium as a secondary messenger
These include muscle contraction, neuronal transmission as in an excitatory synapse, cellular motility (including the movement of flagella and cilia), fertilisation, cell growth or proliferation, learning and memory as with synaptic plasticity, and secretion of saliva. Other biochemical roles of calcium include regulating enzyme activity, permeability of ion channels, activity of ion pumps, and components of the cytoskeleton. Specific signals can trigger a sudden increase in the cytoplasmic Ca2+ level up to 500–1,000 nM by opening channels in the endoplasmic reticulum or the plasma membrane. The most common signaling pathway that increases cytoplasmic calcium concentration is the phospholipase C pathway. Many cell surface receptors, including G protein-coupled receptors and receptor tyrosine kinases activate the phospholipase C (PLC) enzyme. PLC hydrolyses the membrane phospholipid PIP2 to form IP3 and diacylglycerol (DAG), two classical second messengers. DAG activates the protein kinase C enzyme, while IP3 diffuses to the endoplasmic reticulum, binds to its receptor (IP3 receptor), which is a Ca2+ channel, and thus releases Ca2+ from the endoplasmic reticulum. Depletion of calcium from the endoplasmic reticulum will lead to Ca2+ entry from outside the cell by activation of “Store-Operated Channels” (SOCs). This inflowing calcium current that results after stored calcium reserves have been released is referred to as Ca2+-release-activated Ca2+ current (ICRAC). Many of Ca2+-mediated events occur when the released Ca2+ binds to and activates the regulatory protein calmodulin. Calmodulin may activate calcium-calmodulin-dependent protein kinases, or may act directly on other effector proteins. Besides calmodulin, there are many other Ca2+-binding proteins that mediate the biological effects of Ca2+.
resting concentration of Ca2+
The resting concentration of Ca2+ in the cytoplasm is normally maintained in the range of 10–100 nM. To maintain this low concentration, Ca2+ is actively pumped from the cytosol to the extracellular space and into the endoplasmic reticulum (ER), and sometimes in the mitochondria. Certain proteins of the cytoplasm and organelles act as buffers by binding Ca2+. Signaling occurs when the cell is stimulated to release calcium ions (Ca2+) from intracellular stores, and/or when calcium enters the cell through plasma membrane ion channels.
EF hand
a helix-loop-helix structural domain or motif found in a large family of calcium-binding proteins. The EF-hand motif contains a helix-loop-helix topology, much like the spread thumb and forefinger of the human hand, in which the Ca2+ ions are coordinated by ligands within the loop. The EF-hand consists of two alpha helices linked by a short loop region (usually about 12 amino acids) that usually binds calcium ions. EF-hands also appear in each structural domain of the signaling protein calmodulin and in the muscle protein troponin-C. Upon binding to Ca2+, this motif may undergo conformational changes that enable Ca2+-regulated functions as seen in Ca2+ effectors such as calmodulin (CaM) and troponin C (TnC)
Calmodulin (CaM)
a calcium-binding messenger protein expressed in all eukaryotic cells. CaM is a multifunctional intermediate messenger protein that transduces calcium signals by binding calcium ions and then modifying its interactions with various target proteins. CaM mediates many crucial processes such as inflammation, metabolism, apoptosis, smooth muscle contraction, intracellular movement, short-term and long-term memory, and the immune response. CaM is expressed in many cell types and can have different subcellular locations, including the cytoplasm, within organelles, or associated with the plasma or organelle membranes. Many of the proteins that CaM binds are unable to bind calcium themselves, and use CaM as a calcium sensor and signal transducer. CaM can also make use of the calcium stores in the endoplasmic reticulum, and the sarcoplasmic reticulum. CaM can undergo post-translational modifications, such as phosphorylation, acetylation, methylation and proteolytic cleavage, each of which has potential to modulate its actions. It contains four EF-hand motifs, each of which binds a Ca2+ ion. Each Ca2+ coordinated with 5 oxygens (1 backbone, 3 aspartates, 1 glutamate.
Roles of calmodulin
Calmodulin binds to and confers Ca2+ regulation to a large number of other proteins, including ion channels, protein kinases and phosphatases, and cyclic nucleotide phosphodiesterases. The EF-hand motif of calmodulin is found in many other Ca2+ effectors, including parvalbumin (a cellular Ca2+ buffer), calpain (a Ca2+-activated protease) and troponin. There are also many Ca2+-binding motifs that do not resemble C2 domains or EF hands.
C2 domain
a protein structural domain involved in targeting proteins to cell membranes. The typical version (PKC-C2) has a beta-sandwich composed of 8 β-strands that co-ordinates two or three calcium ions, which bind in a cavity formed by the first and final loops of the domain, on the membrane binding face. Many other C2 domain families don’t have calcium binding activity. C2 domains are frequently found coupled to enzymatic domains; for example, the C2 domain in PTEN, brings the phosphatase domain into contact with the membrane where it can dephosphorylate its substrate, phosphatidylinositol (3,4,5)-trisphosphate (PIP3), without removing it from the membrane
stem-cell niche
Stem-cell populations are established in ‘niches’ — specific anatomic locations that regulate how they participate in tissue generation, maintenance and repair. The niche saves stem cells from depletion, while protecting the host from over-exuberant stem-cell proliferation. It constitutes a basic unit of tissue physiology, integrating signals that mediate the balanced response of stem cells to the needs of organisms. Yet the niche may also induce pathologies by imposing aberrant function on stem cells or other targets. The interplay between stem cells and their niche creates the dynamic system necessary for sustaining tissues, and for the ultimate design of stem-cell therapeutics. Cells, matrix glycoproteins and the three-dimensional spaces they form provide ultrastructure for a stem-cell niche. The contact between these elements allows molecular interactions that are critical for regu- lating stem-cell function. Secreted proteins offer a paracrine measure of control, but non-protein components of the local microenviron- ment also affect stem-cell function.
The niche as a target in cancer therapy
If can- cer is organized in a manner similar to normal tissue, with a minor subpopulation of stem cells, an attendant blood supply and a unique microenvironment, there might be a similar dependence of the stem cells on a cancer niche. If the components of this niche could be demonstrated and targeted, it would be of considerable interest to modify the relative support of the stem-like cells of cancer. The idea that the niche might be a druggable target would be extremely appealing as an adjunctive and entirely independent means of targeting malignant cells.
Adult tissue stem cells
First, a population of tissue stem cells should be maintained over long periods of time, often the entire lifetime of the organism (“longevity”). And second, these long-lived stem cell populations should be able to gen- erate the differentiated cell types of the tissue. Most adult stem cells can generate multiple cell types (“multipotency”), yet examples also exist in which they only generate offspring of a single lineage. Stem cells are commonly believed to di- vide very infrequently (“quiescence”), and when this occurs, to generate one rapidly cycling daughter cell, while the other daughter replaces the parent stem cell (“asymmetric cell division”). Most adult stem cells are lineage-restricted (multipotent) and are generally referred to by their tissue origin (mesenchymal stem cell, adipose-derived stem cell, endothelial stem cell, dental pulp stem cell, etc.).
transit-amplifying (TA) cells
The rapidly cycling daughter cells, also called transit-amplifying (TA) cells, are responsible for building tissue mass. TA cells typically undergo a limited number of cell divisions, after which they termi- nally differentiate.
Induced pluripotent stem cells
a type of pluripotent stem cell that can be generated directly from adult cells. Since iPSCs can be derived directly from adult tissues, they not only bypass the need for embryos, but can be made in a patient-matched manner, which means that each individual could have their own pluripotent stem cell line. iPSCs are typically derived by introducing a specific set of pluripotency-associated genes, or “reprogramming factors”, into a given cell type. The original set of reprogramming factors (also dubbed Yamanaka factors) are the genes Oct4 (Pou5f1), Sox2, cMyc, and Klf4. While this combination is most conventional in producing iPSCs, each of the factors can be functionally replaced by related transcription factors, miRNAs, small molecules, or even non-related genes such as lineage specifiers.
Limitations of the transcription factor approach for IPSC
Throughput: the throughput of successfully reprogrammed cells has been incredibly low. Genomic Insertion: genomic integration of the transcription factors limits the utility of the transcription factor approach because of the risk of mutations being inserted into the target cell’s genome. Tumors: another main challenge was mentioned above – some of the reprogramming factors are oncogenes that bring on a potential tumor risk. Inactivation or deletion of the tumor suppressor p53, which is the master regulator of cancer, significantly increases reprogramming efficiency. Incomplete reprogramming: reprogramming also faces the challenge of completeness. This is particularly challenging because the genome-wide epigenetic code must be reformatted to that of the target cell type in order to fully reprogram a cell.
cancer stem cells (CSCs)
are cancer cells (found within tumors or hematological cancers) that possess characteristics associated with normal stem cells, specifically the ability to give rise to all cell types found in a particular cancer sample. CSCs are therefore tumorigenic (tumor-forming), perhaps in contrast to other non-tumorigenic cancer cells. CSCs may generate tumors through the stem cell processes of self-renewal and differentiation into multiple cell types. Such cells are proposed to persist in tumors as a distinct population and cause relapse and metastasis by giving rise to new tumors. Cancer stem cells are also capable of resurrection after morphological and biochemical apoptosis by evoking blebbishield emergency program.[1] Therefore, development of specific therapies targeted at CSCs holds hope for improvement of survival and quality of life of cancer patients, especially for patients with metastatic disease.In brief, CSC can be generated as: mutants in developing stem or progenitor cells, mutants in adult stem cells or adult progenitor cells, or mutant differentiated cells that acquire stem like attributes. Some researchers favor the theory that the cancer stem cell is generated by a mutation in stem cell niche populations during development.
Stem cell niche
refers to a microenvironment where stem cells are found, which interacts with stem cells to regulate cell fate. The word ‘niche’ can be in reference to the in vivo or in vitro stem cell microenvironment. During embryonic development, various niche factors act on embryonic stem cells to alter gene expression, and induce their proliferation or differentiation for the development of the fetus. Within the human body, stem cell niches maintain adult stem cells in a quiescent state, but after tissue injury, the surrounding micro-environment actively signals to stem cells to promote either self-renewal or differentiation to form new tissues. Several factors are important to regulate stem cell characteristics within the niche: cell-cell interactions between stem cells, as well as interactions between stem cells and neighbouring differentiated cells, interactions between stem cells and adhesion molecules, extracellular matrix components, the oxygen tension, growth factors, cytokines, and the physicochemical nature of the environment including the pH, ionic strength (e.g. Ca2+ concentration) and metabolites, like ATP, are also important. The stem cells and niche may induce each other during development and reciprocally signal to maintain each other during adulthood.
Cellular Signal Transduction
Extracellular Signal = Ligand -> Transmembrane Receptors -> Activation of intracellular Signal Transducers -> Modulation of Channels and Enzyme Effectors -> Production of Diffusible Second Messengers -> Regulation of Target Protein Phosphorylation by Kinases and Phosphatses -> specific cellular responsees
agonist
favors the conformation change in the receptor and triggers activation
antagonist
binding blocks receptor activation, usually through competive binding. 50% of all non anti-biotic prescription drugs act through these receptors.
Bordella Pertussis (whooping cough)
PTX: Pertussis Toxin-ADP ribosylates Gαi/o family near the C-terminus of α subunit to lock the G protein heterotrimer in the inactive state by preventing receptor coupling. No activation.
Vibrio cholera (Cholera)
CTX: Cholora Toxin-ADP ribosylates Gαs near the GTP binding site of the α subunit to inhibit GTPase activity thus converting the G protein to the active state even without receptor activation. No deactivation.
Diversity in Signaling through G protein-coupled Receptors
Ligands- different neurotransmitters, hormones and peptides -> GPCRs- bind to different receptor classes & subtypes -> Receptors couple to different G protein transducers -> G proteins regulate different Effector enzymes and channels -> Resulting in production of different Second Messengers -> Protein Phosphorylation- control of different kinases and phosphatases acting on different target substrates -> specific cellular responses
Postganglionic Efferents
Includes those released by Sympathetic Branch- adrenergic transmission at target organs- norepinephrine (NE) released at synapses and epinephrine (adrenaline) released into blood by the adrenal glands and the Parasympathetic branch- muscarinic cholinergic transmission at target organs- acetylcholine (Ach) released at synapses. In general sympathetic and parasympathetic oppose each other in regulating target organ functions/ Sympathetic fight or flight response. Examples in cardiovascular and pulmonary system
Adrenergic Receptors
Include beta1AR and alpha1AR
Muscarinic receptors
includes m2 AchR and m3 AchR
β1ΑΡ
a beta-adrenergic receptor, and also denotes the human gene encoding it. It is a G-protein coupled receptor associated with the Gs heterotrimeric G-protein and is expressed predominantly in cardiac tissue. Gs renders adenylate cyclase activated, resulting in increase of cAMP. cyclic AMP works by activating protein kinase A (PKA, or cAMP-dependent protein kinase). PKA phosphorylates L-type Ca2+ Channels, Ryanodine Receptors (RyR) in SR (Ca2+ release channels) and contractile proteins. Ca2+ influx increases, increased heart rate and contraction. Agonists- Norepinephrine, Epinephrine or Isoproterenol. (antagonists- propanolol or metoprolol)
α1ΑΡ
a G protein-coupled receptor (GPCR) associated with the Gq heterotrimeric G-protein. It consists of three highly homologous subtypes, including α1A-, α1B-, and α1D-adrenergic. Catecholamines like norepinephrine (noradrenaline) and epinephrine (adrenaline) signal through the α1-adrenergic receptor in the central and peripheral nervous systems. α1-Adrenergic receptors are members of the G protein-coupled receptor superfamily. Upon activation, a heterotrimeric G protein, Gq, activates phospholipase C (PLC), which causes an increase in IP3 (by cleaving PIP3) and calcium. This triggers further effects, primarily through the activation of an enzyme Protein Kinase C. This enzyme, as a kinase, functions by phosphorylation of other enzymes causing their activation, or by phosphorylation of certain channels leading to the increase or decrease of electrolyte transfer in or out of the cell. IP3 and DAG second messenger levels increase. IP3 binds to IP3-Receptor in ER and releases Ca2+ DAG-PKC stimulates Ca2+ influx (L-type Channel). Ca2+ stimulates smooth muscle contraction. Peripheral vasconstriction decreases blood flow to skin increasing blood pressure (and also helps shift blood flow to heart, lungs, and skeletal muscle).
Agonists- Norepinephrine, Epinephrine or Phenylephrine
m2-muscarinic cholinergic receptor (m2 AchR)
M2 muscarinic receptors act via a Gi type receptor, which causes a decrease in cAMP in the cell, generally leading to inhibitory-type effects. They appear to serve as autoreceptors. In addition, they modulate muscarinic potassium channels. In the heart, this contributes to a decreased heart rate. They do so by the G bèta gamma subunit of the G protein coupled to M2. This part of the G protein can open K+ channels in the parasympathetic notches in the heart, which causes an outward current of potassium, which slows down the heart rate. The agonist is acetylcholine. parasympathetic > sympathetic
input. cAMP second messenger levels decrease. Protein Kinase A (PKA) activity decreases. L-type and RyR Ca2+ Channels phosphorylation decreases (phosphatases). Ca2+ influx decreases leading to decreased heart contraction . Opening K+ channels leads to Membrane hyperpolarization and decreased excitability. Decreased Ca2+ influx through L-type Ca2+ channels. Decreased heart rate and contraction.
m3 AchR
is coupled to Gq and Stimulates phospholipase C (PLC) Ca2+ and lipid signals. Like the M1 muscarinic receptor, M3 receptors are coupled to G proteins of class Gq, which upregulate phospholipase C and, therefore, inositol trisphosphate and intracellular calcium as a signalling pathway. The calcium function in vertebrates also involves activation of protein kinase C and its effects. Because the M3 receptor is Gq-coupled and mediates an increase in intracellular calcium, it typically causes constriction of smooth muscle, such as that observed during bronchoconstriction. However, with respect to vasculature, activation of M3 on vascular endothelial cells causes increased synthesis of nitric oxide, which diffuses to adjacent vascular smooth muscle cells and causes their relaxation and vasodilation, thereby explaining the paradoxical effect of parasympathomimetics on vascular tone and bronchiolar tone. Indeed, direct stimulation of vascular smooth muscle M3 mediates vasoconstriction in pathologies wherein the vascular endothelium is disrupted. IP3 and DAG second messenger levels increase. IP3 binds to IP3-Receptor in ER and releases Ca2+. DAG-PKC stimulates Ca2+ influx (L-type channel). Ca2+ stimulates smooth muscle contraction, which leads to Bronchoconstriction
phosphodiesterase (PDE)
any enzyme that breaks a phosphodiester bond. The cyclic nucleotide phosphodiesterases comprise a group of enzymes that degrade the phosphodiester bond in the second messenger molecules cAMP and cGMP. They regulate the localization, duration, and amplitude of cyclic nucleotide signaling within subcellular domains. PDEs are therefore important regulators of signal transduction mediated by these second messenger molecules. Inhibitors of PDE can prolong or enhance the effects of physiological processes mediated by cAMP or cGMP by inhibition of their degradation by PDE. Sildenafil (Viagra) is an inhibitor of cGMP-specific phosphodiesterase type 5, which enhances the vasodilatory effects of cGMP in the corpus cavernosum and is used to treat erectile dysfunction.
beta-2 adrenergic receptor
a beta-adrenergic receptor within a cell membrane which reacts with adrenaline (epinephrine) as a hormone or neurotransmitter affecting muscles or organs. his receptor-channel complex is coupled to the Gs G protein, which activates adenylyl cyclase, catalysing the formation of cyclic adenosine monophosphate (cAMP) which then activates protein kinase A, and the counterbalancing phosphatase PP2A. The assembly of the signaling complex provides a mechanism that ensures specific and rapid signaling. A two-state biophysical and molecular model has been proposed to account for the pH and REDOX sensitivity of this and other GPCRs.
