culture issue of postnatal 1 rat cortical neuron culture

[Reinhard]

Dear all,

We're wondering whether anybody had experience in P1 rat cortical culture on MEA? We have tested our system many times with function generator and culture MEAs, the system seems well with a input from function generator, while no signal ever appeared within a cultured MEA. 

We also tried add a droplet of KCl solution to increase the concentration of K+  to 80mM in the MEA, but still no signal appeared in NR.

Since the signal from function generator could be seen, I think the problem may be the cell culture even the morphology seems well. Did anybody meet the same problem?

BTW, the NR we used is 1.1.0.564.

[Riley Zeller-Townson]

Reinhard

If I remember correctly, some folks have cultured P1 cortex using the same techniques demonstrated in the Potter lab Jove article, link below.  

If you are confident that electrically your system is operational, here are some questions:

1) what is the impedance of the MEAs you are using?   NR has a built in impedance measurement capability (if you have the stimulation system rigged up) which can be used to answer this.  If the impedance of a particular electrode is less than 20kohms at 1khz,  it is probably unable to pick up neural  signals. If you  use the impedance measurement in NR,  make sure to calibrate first with a known impedance, such as a test MEA.  If the MEAs you are using are new, this is probably not an issue, however we have found that MEA impedance degrades with use.  

2) what evidence do you have that your cells are alive at the time of recording? 

3) what density do you plate your cells at, and what media do you  keep them in?  the media suggested in the article below was specially formulated to encourage lots of activity.

4) when you record, what temperature are the cells at, and what age?  How long do you let the cells sit in their incubator before you begin recording?  

http://www.jove.com/video/2056/how-to-culture-record-stimulate-neuronal-networks-on-micro-electrode

Hope this helps,  good luck!

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Riley Zeller-Townson

PhD student, Potter lab

Laboratory for Neuroengineering

Georgia Tech/Emory University

Atlanta, GA, USA

[Jon Newman]

Just as general advice,  answering the questions posed by Riley is an excellent starting point for anyone who wants to get recordings from cultured networks.

Jonathan Newman

Postdoctoral Fellow, MIT

http://www.mit.edu/~jpnewman/

[Alessandro]

Dear Reinhard,

if I may add to what already said by Dr. Newman and Riley, in the past I have successfully cultured P1 mouse cortical neuronal cultures (that seem to be notoriously harder to deal with than rat cultures) and carried out MEA recordings with them. I performed the experiments with an MEA-INV-1060 systems and the neurons were plated on 60MEA-200/30iR-Ti-gr dishes. As already mentioned, you are probably having issues with your cultures, rather then with the acquisition system. 

Try culturing your neurons with this medium: http://www.brainbitsllc.com/nbactiv4-for-primary-neuron-cultures/

It is on the pricey side, but it is formulated to increase neuronal activity. Another thing you could try is to plate your neurons at higher cell densities than what recommended for prenatal cultures and this also will help you see more functional activity.

Good luck with your experiments.

Regards,

Alessandro Napoli, PhD

Department of Electrical and Computer Engineering,

Temple University, Philadelphia