Hi Tomasz,

Thank you so much for your suggestion.

I always use voltage stimulation, and for basal synaptic transmission I only recorded pair pulse facilitation. Maybe, I should have done input-output curve and PPF at the same time.

Actually, you pointed some of my worries that "increasing pulse duration will result in activation of more fibers" as well as potential damage to the tissue. However, on the other hand, such increasing duration only happens during stimulation and the duration is the same before and after the stimulation. Ideally, I would like to imagine that the same stimulation, before and after TBS, should stimulate same fibers, although I don't know if that is true.

More important question for me, as a beginner in field recording for LTP, is how to set a proper positive control for LTP itself. Alternatively, what are important factors, for me for pay attention to, in order to get reliable LTP induction in most if not all WT slices?

Thank you for your attention

Fading

Hi Fading,

Tomasz has already raised some good points about the stim duration, so I won't add anything more about that. Something else to consider however, is the method of hippocampal slice preparation for LTP studies. As you may already know, there is a difference in the ability of the dorsal and ventral hippocampal regions to exhibit LTP, with LTP being very much reduced or even absent in the rodent ventral hippocampus compared to dorsal. There are several studies published showing this, for example: http://www.ncbi.nlm.nih.gov/pubmed/10688058

Since you are preparing horizontal hippocampal slices, you are almost certainly using slices from the ventral hippocampus. Perhaps you would have more luck getting consistent LTP in control / WT slices by using slices originating from the dorsal hippocampus. You can prepare these by either making transverse slices (i.e. first dissecting out the hippocampus from the surrounding structures), or by making sagittal slices of the two hemispheres.

Cheers,

Patrick.

http://www.ncbi.nlm.nih.gov/pubmed/10688058

Some interesting points and good tips from the others here. I thought I'd add that there is quite a long history of different labs doubling the pulse width during the induction phase of LTP for short intervals. Usually this i done for either Theta-patterned stimulation of short bursts of high frequency stimuli (i.e. 0.5 sec of 100Hz pulses). In either case, a 0.2 ms pulse for this brief duration is acceptable if you're using a bipolar electrode. Yes you will stimulate more fibers during the induction, but this just helps to create an associative condition where the post synaptic cell is depolarized more during afferent input, enhancing LTP in the fibers you stimulated with lower intensity stimuli. If you do an I/O curve, you should get a leftward shift across stimulus intensities.

Most labs also start off by working at 50% of the maximal response size for a given area. There may be small differences in dorsal/ventral LTP with some paradigms in the CA1 region, but you should keep track of where you are in the hippocampus anyways. My students always keep track of the slice number and usually even quickly draw the shape of the hippocampus. With transverse sections, it becomes quite elongated the more ventral you go. 

Positive controls for LTP are tricky. Really a positive control shows you should get LTP when you expect to, so if you start the day and end the day with slices you can get LTP in, then you're showing it's possible. You could also have an antagonist in place and block LTP initially, and then show you can get LTP when it's washed out. Neither are satisfying positive controls, and your paired-pulse tests and an I/O curve pre and post for every slice is a better way to show you have good slice health for the duration of your recording. 

I hope this helps. We have some reliable ACSF mixtures and stimulus procedures for mice described in the following papers. 

Hi Patrick and Brian,

Thank you both for all insightful points and suggestions. I've been practicing everyday recently using your tips, and it turns out that I can get relatively reliable LTP induction in wild type mouse hippocampal slices. Like you guys mentioned, healthy slices are the key for LTP experiments. 

Cheers.