cpps 325: Inflammatory Pain L15-16

fundamentally, how is pain sensed and sensitivity to it increased

through intracellular signalling

what concept does inflammatory pain signalling illustrate well

multiple pathways converging and cross talking

what is the evolutionary purpose of pain

to alert us that a tissue is being damaged, so we can stop it

aside from a purely molecular signalling component, pain also has an _______ component

emotional: ex) depression amplifies pain

what morphology do sensory neurons take

pseudo unipolar: one branch is the free endings in the tissue, the other runs to the spinal cord

what is the morphology of motor neurons

multipolar

where are the cell bodies of sensory neurons located

the dorsal root ganglia

what is important to realize about the collection of neurons found in the DRG

the ones there are responsible for ALL sensory stimuli: nociception, proprioception, heat, etc.

how do nociceptive sensory neurons tend to look relative to other sensory neurons

nociceptive neurons are smaller and less myelinated

what are nociceptors

the receptors (whether they be GP-coupled, inotropic, etc) that are on the free nerve endings of sensory neurons that respond to noxious stimuli

where in the body are nociceptors found

which body location do we focus on

many places: skin, sheath around muscle, cornea, pulp of teeth

we focus on the skin

while we focus on pain signalling in peripheral neurons, it is important to note there are also other neurons involved

what are they

CNS neurons: those at the level of spinal cord and brain

ie) secondary neurons in the pathway

what are the 2 fundamental responses nociceptors can have in response to signalling molecules

I) channel opening that depolarizes the neutron and fires an AP

II) activation of gene trancription, producing more molecules involved in the pain pathway

what process is key to understand, that occurs as soon as tissue is damaged

damaged cells will release pain signalling molecules

what are some examples of molecules released by tissue damage

NGF, Brady kinin, Protons (drop pH)

give 3 examples of nociceptors and ligands they respond to

TRPV1: Heat, Protons, Capsaicin

TrkA: NGF

BradykininR: Bradykinin

what are examples of cells types that release pain signalling molecules when damaged

T cells, mast cells, macrophages

keratinocytes, fibroblasts, immune cells release NGF

Bradykinin is released by endothelial cells (damaged vessel)

in addition to the pain signalling molecules related by tissue damage, what other players are involved in the nociceptive response

"pain neuropeptides" such as Substance P, and CGRP

these are unique b/c they are released from the nerve fibre itself

in what histological skin layer do nociceptive free nerve endings terminate in

epidermis

(nerve endings that sense pressure are lower, in the dermis)

what kind of receptor is the trkA receptor

RTK

what kind of receptor is the bradykininR

GPCR - linked to Gq

what is one reason NGF has such a crucial role in pain signalling

its RTK links it to the MAPK pathway: this allows it to regulate the expression of molecules such as the pain neuropeptides (SP, CGRP)

so we know that these pain signalling molecules cause an action potential to fire which will be perceived as pain

but they do have other effects - what is the most notable one?

increased SENSITIZATION to pain

also cause vasodilation, leukocyte migration, increased vessel permeability (these things literally are "inflammation")

how is pain sensitization measured

"Von Frey Hairs": small fibres depressing a set amount of force

what is the technical term for being more sensitive to pain "decreased pain tolerance"

Hyperalgesia

what is allodynia

a normally non painful stimulus, becomes painful

what 2 consequences will we see w.r.t pain, after tissue damage

i) Hyperalgesia

ii) the undamaged area adjacent to the original damage, also becomes sensitized

what percentage of sensory neurons contain trkA receptors

~45%

being so crucial to the pain response, the trkA receptor is linked to a lot of pathways (ie. directly via the receptor)

name what 3 pathways these are

i) PI-3K pathway

ii) MAPK pathway

iii) PLC gamma

if you experimentally wanted to identify trkA containing neurons, what would be the best marker to select

CGRP

what would be the effect of a trkA antagonist

abolish the pain sensitized state, for one

but there would be many other effects since NGF is a very widely used molecule in the body

this is why a trkA antagonist is not used therapeutically

what is a better drug target than trkA itself

some of the downstream targets trkA activates

what is a key mechanism that allows sensitization to occur very quickly (ex. 5 minutes after stubbing toe)

Verge lab showed: a decrease in pH = depolarization via TRPV1 = more trkA inserted to membrane

how does trkA insertion happen, mechanistically

via insertion by golgi vesicles

shown b/c adding a drug blocking this pathway resulted in less trkA increase on the membrane

how was it shown that increased number of trkA R's actually led to a heightened response to NGF

An antibody for phospho-trkA intracellular domain showed far greater activation of trkA after exposed to lower pH

what final piece of evidence was used to show lower pH did in fact lead to greater trkA activation and thus increased sensitivity of the sensory neuron

that a downstream target of trkA activity also had its phosphorylation levels increased

describe the cross talk between the BK receptor and trkA receptor, that causes greater depolarization of the sensory neurons

BK couples to Gq, which activates PLC beta, which cleaves the PIP2 gate on the TRPV1 channel

NGF couples to PLC gamma, which does the same

these 2 pathways also release IP3 which causes Ca2+ release from intracellular stores, and increased depol

we have seen that TRPV1 activity can be increased by breaking down the PIP2 gate.

is there another mechanism by which TRPV1 activity can be increased?

Yes: phosphorylation of TRPV1 by PKA or PKC will increase its activity

what structure is important in the increase of TRPV1 activity via phosphorylation

AKAP: bind PKA, PKC, AC, right next to the TRPV1 channel

how is the PLC pathway related to PKC

the breakdown of PIP2 to IP3 activates Ca stores.

Ca2+ activates PKC, as does association with DAG

so then, by what 2 ways does NGF binding to trkA R increase TRPV1 opening

i) via breaking PIP2 gate

ii) via activation PKC which phosphorylates the TRPV1 channel

is there a way to Desensitize the TRPV1 R?

Yes; calmodulin does this

if PKA is to phosphorylate TRPV1, it needs cAMP.

how is cAMP generated in the sensory neuron?

via Gs-coupled R's: which are the receptors for S-P and CGRP!

is TRPV1 the only target of PKA and PKC within the sensory nerve ending?

No: they will also phosphorylate NaV channels: increasing the depolarization

it was mentioned that PI3K was associated with trkA. what does PI3K do?

its activity actually increases the amount of trkA R's being inserted on the membrane!

so you can regulate the activity, and the quantity of the R's

what is a second role of CGRP and SP ligands besides increase cAMP in the neuronal ending

they will diffuse to cells such as mast cells and cause further pain signalling molecule release

what is a third target of PKC regarding the sensory nerve ending

an ATP- activated nonselective cation channel known as P2X3 = depolarization