When it comes to the induction of LTP, people report different
kinetics. I don't think you're problem is the flow rate. 2-3 mL/min is
fine. I think the problem is the bubbles. You're causing electrolytic
lessons. The easiest way to stop the bubbles is to use a glass
microelectrode (500kOhm-2MOhm) filled with aCSF as your stimulating
electrode. Your pulses also seem quite strong (and long). You should
be able to evoke pretty good fEPSPs without resorting to such extreme
stimulus strengths. Finally, 100Hz for 1 second? Have you thought of
switching to a theta burst type paradigm? Most people report stronger
and more stable LTP with theta burst type stimuli.
On Oct 23, 1:47 am, Ana Ruiz Nuño <anaruizn... from yahoo.es> wrote:
> To members to Neur-sci,
>>> I am setting up the LTP in mouse hippocampal slices (350 microns) in a submerged chamber. I get the LTP after 30 min but I think I am still having problems with perfusion because I don't get the typical kinetics as you can see in the attachment.
> Some authors get good kinetics in mouse with 2-3ml/min as I got. Others use faster (5-6ml/min) ones, but sometimes it is not very easy to get an equilibrium between perfusion and stability.
> There is another problem with the mouse slice. Although I place the stimulus and recording within 400-500 microms apart, sometimes you get bubbles after the tetanic stimulation and it takes time till dissapear and get the real effect.
>> I am stimulating and recording in stratum radiatum of the CA1 region. I try to be consistent and the stimulating microelectrode was positioned in the ipsilateral Schaffer collateral-commissural pathway of CA3 area and the recording microelectrode was placed within the stratum radiatum of CA1 area 300-600 microns apart. Synaptic responses were evoked by constant stimulus pulses (0.2 ms, 0.3-2 mA) delivered by bipolar Ni/Cr stimulating electrode.At the beginning of each experiment, test pulses that evoke 40-50% of the maximum EPSP slope.Tetanic stimulation (TS) was a 1-s 100-Hz pulse for induction of LTP. The amount of LTP was quantified 30 min after the train, and was expressed as percent increases in the slope of field EPSPs. Recording chamber at 3 ml/min at 30-32ºC. I also put a piece of lense paper under the slice.
>> Any suggestions for improving recordings?. Do you think is a reliable recording for publication?. I attach a typical recording.
>> Thank you for your time in this matter