Neurobiology H

These two peptides come from the same precursor protein, proopiomelanocortin (POMC).
Melanotropin is found in the N-terminal of corticotropin.

pars intermedia

pars distalis.

*pars intermedia in mammals is only some layers thick.
*research of neuroendocrine integration
in nijmegen.

the regulation of skin color during the process of background adaptation

dispersion of pigment, thus leading to black color.

*On white background the pigment in the melanophores is fully contracted around the nucleus (termed perinucular position);
*Reflected light leads to an activation of the neurons and thus a release of dopamine in the pars intermedia; dopamine inhibits the release of MSH and thus the animal turns white.


*On black background, under the influence of MSH, the pigment becomes fully dispersed throughout the melanophore; this has been designated melanophore index of 5.
*no reflected light > no dopamine (inhibition of dopaminergic neurons) > no MSH inhibition > black

the reflected light overrides the non-reflected light to stimulate the dopaminergic neuron.

to determine of neurotransmitters/neuropeptides are present in pars intermedia or nervosa.
*dopamine is present in nerve terminals in the pars intermedia and showed that the cell bodies for these dopaminergic neurons were in the suprachiasmatic nucleus (SC) of the hypothalamus.

*Do factors(dopamine) effects MSH secretion?

*separation of hypofyse lobs
*PI+PN
*PD
*in saline solution
*introductions of factors, measuring MSH activity with radioimmunoassay.

*dopamine inhibits MSH secretion from the Xenopus melanotrope cell by acting on a dopamine D2 receptor.

GABA,dopamine and a neuropeptide called neuropeptide Y (NPY)

dopamine and NPY are cosequestered in dense core vesicles, while GABA is found in the nerve terminals in electron lucent vesicles.

NPY inhibits MSH secretion through a NPY Y1 receptor while GABA, which also inhibits secretion, works through both a GABAa receptor and a GABAb receptor.

*Superfusion studies showed that both CRH and TRH stimulate secretion of MSH.
*Presumably these stimulatory factors would diffuse from the pars nervosa to the pars intermedia to regulate MSH secretion.

Yes, they stimulate there the release of MSH.

RC neurons: 5-HT , LC neurons: noradrenaline

*Ach, through M1 receptors.

*ATP > P2X,P2Y receptors.
*Ca2+ > Ca sensing receptors.(G-protein coupled receptor).

each regulatory factor seems to have its own properties.

* Ca2+ oscillation (dynamic imaging) drive vesicle secretion (amperiometry)
*AC > cyclic AMP > PKA > membraan potential maschinery (MPo) > Ca2+ oscillations> influx Ca2+ > exocytosis and induce Ca2+ release mechanism that generated a self-propagating wave of Ca2+ that went through the cytosol and entered the nucleus(POMC gene expression).
*Gi linked receptors have a direct action on MPo via the beta/gamma subunit of G protein.

*Na+, K+ and Ca2+

BDNF induceert late LTP

melantrope cells are twice as big in animals on a black background compared to white background adapted animals.
*black animals > reach in RER and display more POMC expression.

*Individual melanotorpe cells display many different patterns of Ca2+ signaling
*Some of these are silent and other, while all displaying Ca2+ oscillations, display very different patterns in their oscillations.
*oscillations in step, no step wise fassion.
*each step = opening of ca channel
*white - silent and no-step patterns are dominant
*black - steppers far of the majority.

*black - higher expression of POMP
- precursor convertase gene involved in POMC processing.
-7B2 which is a caparone (carrier) for the PCs through the regulated secretory pathway
-BDNF
*white: - higher NPY Y1 receptor expression
- higher expression of the stimulatory receptors for TRH and CRH.

*Perhaps the cells are sensitizing themselves so that they can quickly respond if they come into a situation (e.g. black background) where a release of alfa-MSH is called for.
*Alternatively, perhaps the mRNA is not transcribed but rather forms a pool of mRNA available for quick translation if the animal is put on black background.

*melanotrope cell itself has cyclic-AMP and Ca2+ as intracellular message molecules
*yet the promoter region of the POMC gene contains no cyclic-AMP responsive elements (CREs), nor does it contain any Ca2+ responsive elements (CaREs).
*or Fos-c of BDNF is involved

1.BDNF is produced by the Xenopus melanotrope cell
2.that it is cosequestered with alfa-MSH (and thus likely co-released with the melanotropin)
3.that the melanotrope cell expresses the BDNF TrkB receptor
4.that BDNF stimulates the biosynthesis of POMC (production of radiolabelled prohormone in radiolabeling experiments).

*six promoter specific 5‟exons
*each of which can become spliced to a single 3‟ protein coding exon to generate six unique BDNF transcripts
*pre-proBDNF coding exon VII has its own upstream promoter to generate a 5‟ extended transcript (transcript VII-5‟ext).
*the expression of the various promoter-specific BDNF transcripts in Xenopus is tissue specific.
*

transcript IV (black animals)and transcript VII-5‟ext. (black and white animals)

Promoter IV contains 2 CaRE, 1 CRE and a DRE site (binding site for the Ca2+ regulated transcription factor DREAM ).
The promoter region of VII-5‟ext contains none of these responsive elements.

*transcript IV as a regulated transcript and transcript VII-5‟ext as a constitutively expressed transcript.

*The results show that BDNF is clearly up-regulated before POMC, which is in keeping with the idea that BDNF would stimulate POMC gene expression.
*In these same experiments we examined c-Fos expression; the results leave little doubt that c-Fos acts as an immediate early gene in the melanotrope cell in animals adapting to a black background

*PCR to study expression of “growth promoting genes” known to be regulated by BDNF in the CNS.
*The other approach is to use microarrays to determine which genes are being turned on during black background adaptation

Ras

If a cells want a highly efficient production of BDNF then it express a transcript with few uAUGs.

*AUG before the AUG site coding for the protein.
*uORFs makes the translation process less efficient.

Phosphorylation of this factor (by one of the kinases involved in the regulation of the translation complex) inactivates this factor and causes the scanning ribosome to skip over uORFs in a process called leaky scanning.

give ribosomes direct access (without scanning) to the authentic AUG(thus bypassing all the uORFs

block cap-dependent translation and see if the mRNA is still translated…if it is then the mRNA likely possesses an IRES).

*when cell expirience stress
*the mRNA for many of the proteins that are responsible for cell survival have been shown to possess IRESs.

*in the brain BDNF is known to protect neurons from damage and help repair cell damage
*The idea is that during the LTP the synapse switches from a system favoring Cap-dependent translation to a system favoring IRES-dependent translation.
Thus mRNAs possessing IRESs, which before the LTP was quiescent, suddenly are transcribed, leading to a rapid production of proteins involved in synaptic strengthening.