The First Author Forum ION LiYan

The First Author Forum ION LiYan

Santiago Ramón y Cajal

Axon Guidance: an intriguing question from 100 years ago

“From the functional point of view the growth cone may be regarded as a sort of club or battering ram, endowed with exquisite chemical sensitivity, with rapid ameboid movements, and with certain impulsive force, thanks to which it is able to proceed forward and overcome obstacles met in its way, forcing cellular interstices until it arrives at its destination”

Chemotaxis mechanism for axon guidance

Adapted from Nobel Lecture 1906,


Ross Granville Harrison

J. Exp. Zool., 9:787–846 (1910).

1. First observed growth cone movement of neurons growing in tissue culture

2. Contact guidance mechanism


The model systems of studying the axon guidance

In vivo

Vertebrate ventral spinal cord

Optic chiasm

Drosophila ventral nerve cord

In vitro

Explant culture

Turning assay



vertebrate ventral spinal cord

In vivo

Cell, Vol. 87, 1001–1014, December 13, 1996,



In vivo

Optic chiasm

Cell, Vol. 119, 567–578, November 12, 2004,



In vivo

Nat Neurosci. 2005 Feb;8(2):156-63, Jan 16, 2005

Drosophila ventral nerve cord



In vitro

Explant culture

Cell, Vol. 96, 807–818, March 19, 1999,



In vitro

Turning assay


Why did we choose cerebellar granule cells?


How did I modify the turning assay?

New information

Collaboration


Basic question in the axon guidance

1. Finding the new guidance cues

2. Mechanism of axon guidance induced by guidance cues

Ca2+ signal

Cytoskeletal dynamics


How to find the question?

1. Read the text book

Repeat the classic experiments with the new technique to find the novel and undiscovered phenomena

Directly touch the key problems undiscovered and unsolved by others in the field

2. Read the newest review in the field

Cajal1891 Nature,Vol

. 399, 6

May, 1999


Why BDNF?Mice lacking brain-derived neurotrophic factor develop with sensory deficitsErnfors P, Lee KF, Jaenisch R.Nature. 1994 Mar 10;368(6467):147-50

cAMP-induced switching in turning direction of nerve growth conesSong HJ, Ming GL, Poo MMNature. 1997 Jul 17;388(6639):275-9

BDNF stimulates migration of cerebellar granule cells.Borghesani PR, Peyrin JM, Klein R, Rubin J, Carter AR, Schwartz PM, Luster A, Corfas G, Segal RADevelopment. 2002 Mar;129(6):1435-42

2003 SARS


Turning response induced byBDNF is dependent on bothintracellular Ca2+ and extracellular Ca2+

How to enter the growth cone?


Calcium signalling in the guidance of nerve growth by netrin-1

Kyonsoo Hong*², Makoto Nishiyama*², John Henley*,Marc Tessier-Lavigne³ & Mu-ming Poo*

NATURE |VOL 403 | 6 JANUARY 2000

The netrin-1-induced turning response depends on Ca2+ influx through plasma membrane L-type Ca2+ channels, as well as Ca2+- induced Ca2+ release from cytoplasmic stores.


L-Type Ca2+ Channel?

Voltage-dependent Ca2+ Channels (VDCCs)


Neurotrophin-evoked depolarization requires the sodium channel Na(V)1.9.

Blum R, Kafitz KW, Konnerth A.

Nature. 2002 Oct 17;419(6908):687-93.

Neurotrophin-evoked rapid excitation through TrkB receptors

Kafitz KW, Rose CR, Thoenen H, Konnerth A.

Nature. 1999 Oct 28;401(6756):918-21


Hypothesis I:

BDNF Nav1.9Channel

Depolarize membrane VDCCs Ca2+ entry


Our Study


Ca2+ dependent chemoattraction induced by BDNF


Novel Ca2+ entery mechenisam in the turning induced by BDNF

Activation of a TRPC3-Dependent Cation Current through the Neurotrophin BDNF

Hong-Sheng Li, Xian-Zhong Shawn Xu, and Craig Montell*

Neuron, Vol. 24, 261–273, September, 1999,

Hypothesis II:

BDNF TRP Channel Ca2+ entry



TRP channels involved in the turning induced by BDNF


TRP channels mediated Ca2+ entery in the turning induced by BDNF



Choice

Breakthrough Work: Gating mechanism

Solid Work: Examination Subfamily

The expression of TRPC channels in the cerebellar granule cells


TRPC3/6 channels were essential for the BDNF- induced attraction



TRPC3/6 channels were essential for the BDNF- induced Ca2+ signal


Model

Nature, Vol. 434, 14 April 2005


How to finish your project efficiently?

1. Understand the question which you are working on

2. Modify your hypothesis based on your data

3. Work independently

4. Push your PI instead of being pushed

choice


My strategy to find a project in the lab

1. Based on the technique in the lab

2. Hypothesis driven

3. Used and developed new technique based on the experiments


The better way for junior students to begin the project

1. Think about the big problem unsolved in your field

2. Peer the ongoing project and research interest in your lab

3. Open your mind

4. Accumulate the experimental experience

5. Analyze your data in time


It is harmful to wait for your PI arranging everything for you!


The most important thing for us

Interest

Persistence


Thanks!

Documents

The First Author Forum ION LiYan