Intracellular Calcium Homeostasis

Question 1

What is the typical resting Ca concentration in the cytosol?

Answer 1

50-100nM

Question 2

How do cells keep IC calcium low?

Answer 2

PM Ca-ATPase, ER-Ca-ATPase, mitochondrial high capacity(low affinity) pump - when IC is very high, existence of Ca binding molecules in cytoplasm reducing free Ca

Question 3

What are the pumps that exist on the ER and mitochondrial membrane for keeping Ca low?

Answer 3

Ca-ATPase, NCX, Ca-binding molecules

Question 4

What are the pathways for IC calcium increase?

Answer 4

VGCC, ROCC, NCX, Ca-ATPawe of SER and PM, Ca release channels on SR and ER

Question 5

What does the removal of K via of BK channels do to IC Ca?

Answer 5

causes VGCC and ROCC to close

Question 6

How can calcium be released from Internal stores?

Answer 6

ligand binding to GPCR - Gq - PLCB1 catalysis hydrolysis of PIP2 which produces IP3 and DAG - IP3 release Ca from ER

Question 7

What is the effect of lithium on PIP2 metabolism?

Answer 7

inhibits enzymes involved in the recycling of phosphoinositols which blocks the regeneration of PIP2 and therefore depletes PIP2
overactivity of PIP2 can result in mania so lithium can treat this but may affect other organ functions

Question 8

What is the main subtype of IP3R in the brain?

Answer 8

IP3R1

Question 9

How is the depletion of IC calcium stores sensed?

Answer 9

STIM proteins

Question 10

What channels are activated by STIM proteins to stimulate store refilling?

Answer 10

ORAI proteins

Question 11

Which drugs can be used to prevent Ca requestration into stores?

Answer 11

thapsigarin or CPA

Question 12

What is the difference between STIM 1 and STIM 2?

Answer 12

STIM1 is a sensor of store depletion where STIM2 senses basal ER levels

Question 13

How are ORAI proteins stimulated?

Answer 13

the C domain of the ORAI is stimulated by STIM. EF hand senses the availability of Ca, low affinity so high Ca will bind when store full and releases when depleted
STIM releases Ca when depleted and initiates a process leading to CRAC channel activation
mutants of EF hand show this is its function

Question 14

Where does STIM1 bind?

Answer 14

the microtubule of the end binding protein 1

Question 15

How does the cell respond to sudden changes in energy demand?

Answer 15

by sensing an increase in mitochondrial Ca

Question 16

What happens when the mitochondria is overloaded with Ca?

Answer 16

causes the opening of a high conductance channel, the mitochondrial permeability transition pore (PTP) leading to collapse of proton gradient and ATP production, loss of mitochondrial membrane integrity and some forms of cell death

Question 17

What are the main pathways for Ca entry and exit form the mitochondria?

Answer 17

Mitochondrial calcium uniporter - low conductance, low affinity
mitochondrial NCX
mHCX
mPTP - may activate Ca dependent proteases wsuch as calpaan to initiate cell death

Question 18

Why are mitochondria placed close to ER/SR?

Answer 18

can sense local intracellular Ca conc. and serve as sensors of cellular Ca signals MFN2 helps form mitochondria ER contacts
MAMs are where the ER and mitochondrial Ca channels interact

Question 19

how might mitochondria be positioned in a neuron?

Answer 19

they may be positioned near Ca conduction channels to ensure that the accumulation of Ca does not cause a cytosolic Ca wave

Question 20

What are some of the spatial and temporal aspects of Ca signalling?

Answer 20

transient Ca may be localised and specifically regulate only nearby target e.g. NT release
Ca ions ca propagate as a wave throughout a cell e.g. to communicate signals to the nucleus i.e. at fertilisation
(if sustained this can cause neurotoxicity so waves are infrequent)

Question 21

What are the two binding sites on the IP3R?

Answer 21

one high affinity binding site for Ca to stimulate further release and one low affinity binding site to reduce Ca release in high concentrations

Question 22

How does Ca affect CaM?

Answer 22

has 4 Ca binding sites and the binding of Ca increases its affinity for other target enzymes

Question 23

How does Ca-CaM modulate Ca-CaM dependent Kinase II?

Answer 23

Ca-CaM removes the auto inhibitory domain for the catalytic site resulting in autophosphorylation and activation of the AMPAR increasing its current and adding new AMPAR to the synapse
Constituitively active before but Ca-CaM increases activation
important in LTP and concentrated in post-synaptic density
in low calcium conditions calmodulin dissociates after 10s

Question 24

What is the role of Calcinuerin in LTD?

Answer 24

Ca-CaM activates Calciuneurin which is a dependent phosphatase and regulates a phosphatase, phosphatase 1.
inhibition of Calcineurin blocks induction of LTD
LTD results from removal of AMPAR by endocytosises

Question 25

What are some methods of measuring CA?

Answer 25

radioactive labels - not great as no spatial/temporal info
ion-sensitive micro electrodes - no spatial info
indirect electrophysiology - Ca dependent currents
NMR spectroscopy
Ion-sensitive dyes