PSC2002/L05 NaCl Absorption Flashcards
Give 4 pathways in which NaCl and water pas through the epithelium.
Passive facilitative entry of Na+ across apical membrane
Active exit of Na+ across basolateral (serosal) membrane
Paracellular diffusion of Cl- through tight junctions
Osmotically driven water absorption
Describe the pump-leak model of NaCl and fluid absorption in epithelial cells. (4)
Na influx via EnaC (G(Na))
Na+ actively pumped across basolateral membrane via NA+/K+ ATPase
Causes paracellular transport of Cl- via tight junctions (TJ) to maintain electroneutrality
Increases NaCl concentration on basolateral side & drives osmotic movement through AQPs or TJs
What is the role of ENaC in:
a) the kidney
b) the lungs
c) colon
d) sweat glands?
a) Na+ retention, control of whole-body Na+ and water balance, BP
b) Na+ and water reabsorption, control of airway surface liquid (ASL) and alveolar lining fluid (ALF)
c) Na+ and water reabsorption from diet
d) Na+ retention, reabsorption of Na+ by sweat ducts, not followed by water, produces a hypotonic sweat secretion
What is the functional channel of ENaC?
Hetrotrimer of 3 (aBy) subunits coded by 3 genes
How many transmembrane domains does ENaC have?
2, forming the pore of the channel
Give a potent specific inhibitor of ENaC.
Amiloride
What are the long extracellular loops the site for on ENaC?
CAPs and SPLUNC1 regulation
What is the PY-motif in C-term site for in ENaC?
Ubiquitination
Describe the role of ENaC in the kidney.
Determines final urinary salt composition through natriuretic hormones e.g., aldosterone
How does aldosterone affect ENaC function?
Changes ENaC function in aldosterone-sensitive distal nephron (ASDN)
Stimulates Na+ reabsorption through ENaC by principle cells in ASDN
Increased NaCl and H2O absorption, increased blood volume and BP
Where is the ASDN (aldosterone-sensitive distal nephron)?
Last third of DCT, CNT and cortical collecting duct (CCT)
Describe the action of aldosterone on principal cells on the distal nephron. (3)
Binds to cytosolic mineralocorticoid receptor (MR)
Binds to genes with HREs in nucleus
SGK levels increase with 1 hr
What does aldosterone increase? (4)
Surface ENaC levels 2-5 fold
Na+/K+ ATPase activity
ATP supply to support increased Na+/K+ ATPase activity
K+ excretion across apical membrane via ROMK
What is the number of channels in the apical membrane controlled by?
Rate of insertion versus rate of retrieval/degradation
How does aldosterone increase number of ENaC channels?
By decreasing rate of retrieval
What is the retrieval and degradation of ENaC regulated by? (3)
Ubiquitin ligase NEDD4-2
Binds to PY motif of a, B or yENaC and adds ubiquitin group to lysine residue in N-term of ENaC subunits
Signals internalisation of ENaC followed by degradation
How does aldosterone prevent ubiquitination of ENaC? (3)
Stimulates production of serum and glucocorticoid regulated kinase 1 (SGK1)
SGK1 phosphorylates NEDD4-2 and prevents binding to ENaC
Inhibits ENaC ubiquitination
How is hypertension caused by dysregulated ENaC?
- Excess aldosterone or mineralocorticoid action of other steroids
Aldosteronism
Glucocorticoid-remediable aldosteronism (GRA)
Apparent mineralocorticoid excess (AME) - Liddle’s syndrome - gain of function in ENaC
Mutations in cytoplasmic C-terminal region of B- or yENaC
Describe Liddle’s Syndrome.
Gain of function mutations in ENaC
Rare, autosomal dominant genetic disease caused by mutations in genes encoding 3 subunits SCNN1A/B/G
Leads to cytoplasmic region changes in ENaC subunits
Prevents NEDD4-2 binding and increased number of ENaC causing hypertension
Leads to low blood K+ (hypokalaemia)
Describe how Liddle’s Syndrome causes enhanced K+ excretion.
Enhanced ENaC function depolarises apical m.p.
Electrical gradient for K+ excretion through ROMK increased
Excessive loss of K+ in urine leading to low blood K+
Give a drug which helps restore normal blood K+ and explain how this works.
Amiloride
Inhibitor of ENaC
Hyperpolarises apical m.p.
Reduces K+ loss and lowers BP
(K+ sparing diuretic)
Where is ENaC expressed in the lungs? (2)
Apical membrane of surface epithelial and alveolar type II cells
Describe the role of ENaC in the lungs.
Regulates ASL volume in conducting airways crucial for innate defence of lungs via mucociliary clearance
Regulates ALF volume essential for efficient gas transfer
What is the effect of channel-activating proteases (CAPs) on ENaC in conducting airways?
Increase ENaC activity via cleavage of external loops of a and y ENaC
What is the effect of anti-proteases on ENaC in conducting airways? Give an example.
Reduce ENaC activity by inhibiting proteases
E.g., aprotinin
What is the effect of SPLUNC1 on ENaC in conducting airways?
Reduces ENaC function
Secreted into ASL and binds to ENaC
Protects from internalisation and CAP activation
What is the effect of ATP on ENaC activity in the lungs?
Reduces ENaC activity
Describe the experimental evidence that the protease Cathepsin B (CTSB) regulates ENaC activity in primary cultures of human airway epithelial cells. (3)
Basal cell isolation and expansion from lungs
Differentiation at an air-liquid interface
Pseudostratified single epithelial layer
Inhibiting cathepsin B reduces fluid absorption across monolayers of primary human airway cells
