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Physiology of saliva
DENT 5302
Topics in Dental BiochemistryDr. Joel Rudney
Foundation knowledge
DENT 5315 Oral Histology Dr. Koutlas’ salivary gland lectures
Ten Cate’s Oral Histology Chapter on Salivary Glands
NSCI 6110 Neuroscience for Dental Students Material on neurotransmitters, signal transduction
PHSL 6051 Physiology for Dental Students Material on water transport, signal transduction Future foundation for gastrointestinal and kidney
Innervation of stimulation
Dual autonomic innervation of salivary glands Parasympathetic - secretion of water and ions Sympathetic - protein secretion Both act simultaneously and synergistically
Mediated by G-protein coupled receptors Parasympathetic - M3 muscarinic receptors
Minor players - neuropeptide; nucleotide receptors• VIP, Substance P, nucleotides, etc.
Sympathetic - 2 adrenergic receptorsMinor players - adrenergic receptors
Two different signal transduction pathways
Muscarinic messages
http://www.liv.ac.uk/~petesmif/teaching/1bds_mb/p4/15.gif
The Phospholipase C - IP3 pathway sends the message
Intracellular (and extracellular) Ca2+ flux is a major effector
Adrenergic messages
http://www.liv.ac.uk/~petesmif/teaching/1bds_mb/p4/16.gif
The adenylate cyclase - cAMP pathway sends the message
Effectors are activated by a phophorylation cascade
(Noradrenaline)
Water/electrolyte secretion Water secretion is driven by osmotic changes
Mediated by ionic fluxesFrom basolateral surfaces to the apex (lumen)
Involves ion pumps and channels Basolateral
Na+-K+-ATPase Ca2+ activated K+ channelNa+-K+-2Cl--cotransporter (NKCCl)Na+-H+ exchangerCl-- HCO3
- exchanger, plus Carbonic anhydrase
LumenalCa2+ activated Cl- channelHCO3
- channel (Ca2+ activated?) , plus Carbonic anhydrase
Alternative mechanisms
Na+-K+-ATPase
Ca2+ activated K+ channel
Na+-K+-2Cl--cotransporter
Ca2+ activated Cl- channel
Na+-H+ exchanger
Cl-- HCO3- exchanger
Carbonic anhydrase
Na+-H+ exchanger
HCO3- channel
Carbonic anhydrase
Adapted from Turner and Sugiya, Oral Dis. 2:3-11, 2002
Newly-discovered components
Channels for extracellular Ca2+ in basolateral membrane Initiate Ca2+ flux that activates other ion channels hTrp1, others?
How does water cross the apical membrane? Aquaporin family of water channels
Found in many organ systemsSalivary aquaporin is Aqp5
Ca2+ activated, open to let water outIonic flux pulls the water out - during stimulation
Low level of activation in resting state??
Validation of the model
http://wwwdir.nidcr.nih.gov/dirweb/common/sps.jpgGresz et al. Am. J. Phsiol. 287: G151-G161, 2004
Apical Aquaporin 5
Basolateral Na+-K+-ATPase
Ductal reabsorption
Saliva entering the lumen is isotonic Saliva entering the mouth is hypotonic Reabsorption of Na+ and Cl- by striated duct cells
Similar to distal tubules of kidneys Ion pumps and channels
LumenalNa+-H+ exchangerCl-- HCO3
- exchangerHCO3
- channelNa+-K+ exchangerNa+-Cl--cotransporter
BasolateralNa+-K+-ATPaseCl- channel
Striated duct cell
Cl-
3 Na+
2 K+ATP
Cl-Na+
Na+
Na+
K+
H+
Cl-
HCO3-
HCO3-
Lumen Interstitium
Nucleus
MitochondriaBasolateral membrane folds
Carbonic anhydrase
Clinical significance
Many points for drugs to interfere with water secretion Receptors, signal transduction, ion pumps/channels May explain why xerostomia is a widespread side effect
The M3 receptor is a key point Autoantibodies to M3 occur in some Sjogren’s patients
Sjogren’s etiology and pathogenesis is very complex Agonists can be useful in profound xerostomia treatment
Pilocarpine and CevimilineRequires some remaining functional tissue
Anti-cholinergicsMost likely to induce xerostomia as a side effect
Research “in the pipeline”
Can we repair damaged salivary glands? Gene therapy approach
Use viruses to transfect genes into host cells Infusion into ducts
Ducts are best preserved in Sjogren’s/radiation Transfect aquaporin into rat duct cells
Not normally present in duct cellsTransfection increased salivary flowShort-term effect, and only replaces waterWould need to replace many genes for full repair
Tissue engineering
May have more potential in the long run Step 1: Create a biocompatible scaffold
Must have a duct-like structure Step 2: Seed with cells
Engineer cells to function like secretory/duct cells OR Use stem cells and induce differentiation
Step 3: Implant into a patient Must induce vascularization and innervation Must suppress rejection or use compatible cells Will it make saliva??
Protein secretion
A parallel process to water/ion secretion Both occur side by side in the same secretory cell There is complex cross-talk between pathways
Classic exocytosis pathway Endoplasmic reticulum - translation, glycosylation Golgi - more extensive glycosylation Condensing vacuole - packaging, condensation Immature granule - sorting, major branching point Secretory granule - protein storage -adrenergic stimulation
Docking, membrane fusion, exocytosis
Classic exocytosis
Immediate response to NA:Docking and fusion of preformed granulesRelease of contents
Long-term response to NA:TranscriptionTranslationGlycosylationNew granuleshttp://www.liv.ac.uk/~petesmif/teaching/1bds_mb/p4/14.gif
(Noradrenaline)
Secretory granules
Complex internal structure Multiple types of proteins, compacted and folded
Membrane proteins that mediate docking and fusion V(esicle)-SNARES on granule membranes T(arget)-SNARES on inner side of cell apical membrane A Ca2+ -dependent process
Example of cross talk between pathways
The other protein pathways Constitutive-like pathway
Branches off from immature granules Proteins carried in vesicles to apex - fuse and open Always active - no stimulation required
Minor regulated pathway Branches off from immature granules Proteins carried in vesicles to apex - fuse and open Triggered by low levels of M3 cholinergic agonists Vesicle membranes contain t-SNARES for granules
Both are sources of proteins in basal and resting secretions Vesicle contents are different from granule contents Explains different protein composition after stimulation
Cross-talk is essential
Castle, A. M. et al. J Cell Sci 2002;115:2963-2973
Cholinergic agonist - very low doseConstitutive-like and Minor Regulated only
Cholinergic agonist - low doseConstitutive-like and Minor Regulated with occasional granule docking
Adrenergic agonist - standard doseConstitutive-like and Minor Regulated plus Classic Exocytosis
Adrenergic agonist - standard doseCholinergic agonist - low dose Constitutive-like and Minor Regulated plus synergistic Classic Exocytosis
Supplemental Reading
Turner RJ, Sugiya H (2002). Understanding salivary fluid and protein secretion. Oral Diseases 8:3-11.