This video demonstrates a procedure to quantify four poor dexterity during food handling. In rodents, each animal is exposed to short lengths of uncooked pasta in first the home cage, and then the testing environment. In the testing environment, the animal's fopo movements are videotaped During past eating, the adjustments made with each fopo are identified and countered during slow motion video playback.
During the video analysis, atypical handling behaviors are identified. Identified results show changes in four poor dexterity after brain damage or during aging through the quantification of movements used to manipulate food during eating. Hi, I'm Kelly Tenet from the laboratory of Teresa Jones in the Department of Psychology at the University of Texas at Austin, And I'm RNAi also from the Jones Lab.
Today we're going to show you procedures for the Vermicelli Handling Test in rats and the CAPI handling test in mice. We use these procedures in our laboratory to study how Dexter's four P use during food handling changes, after brain damage or during aging. So let's get started.
To prepare the pasta pieces for testing, use an ultra fine tipped marker to Mark DeCheco brand, CAPI and Skinner brand vermicelli at specific intervals with a Cardstock template. For capal, use 0.86 centimeter increments, and for vermicelli use 1.75 centimeter increments. This facilitates visualization of the movement of the pasta strand.
During eating, using a razor blade cut capi to 2.6 centimeter lengths for use with mice and for rats cut vermicelli to seven centimeter lengths. Because pasta brands vary in texture and diameter, which can influence handling manner, it is recommended that a single source of pasta be used for repeated testing to overcome neophobic responses and to establish skill in handling, expose the animals to the past pieces in their home cage in the weeks prior to testing. Five to 10 days of exposure with about four pieces per animal, each time is sufficient.
Even habituated animals may become reticent to eat in front of the camera. For this reason, place the camera in front of the testing area during one or more habitation sessions. Once the animals have been exposed to the pasta pieces, place them in the testing environment and habituate them to eating there.
Generally, animals are readily eating the pasta in this environment. After three to five eating sessions with preparations complete, let's see how to administer pastor handling trials in brats and mice. Test sessions consist of sequential trials with one past apiece eaten per trial.
A session typically consists of four to five trials per rat and three to four trials per mouse. Rats typically sit upright on their haunches to eat pasta, and the pulls are most easily viewed. With a camera positioned at approximately chamber floor level with a slight upward angle, mice tend to bend forward and hold the pasta at an angle.
Thus, a testing chamber with an open floor is placed on top of a mirror. The camera is placed at a slightly higher level than the floor and angled downward to focus on the reflection of the paws in the mirror. Before initiating the videotaping place a piece of pasta into the testing chamber.
To initiate eating and to move the animal into an appropriate camera view, the animal must face the camera and the camera must be sufficiently zoomed to 10 to 12 x. In order to view discrete adjustments in both four paws and all digits, the knuckles of both paws should be visible as soon as the animal and camera and position initiate the first trial drop or dangle a piece of pasta directly in front of the animal and begin recording. Prior to eating onset, each bite of the pasta piece is audible.
So audio recording is useful for verifying eating. Continue with the remaining trials. In the same way it is best to minimize delay between trials so that the animals do not move away from the optimal viewing position.
Now let's see how to perform quantitative video analysis of four poor adjustments and eating time. The analysis of four poor adjustments during eating is a simple quantitative measure that is sensitive to lateralized impairments after CNS injury in rats and to age related changes in mice during analysis, video playback is slowed to approximately 50%of real time, or frame by frame advance is used as needed to identify adjustments. The quantified portion of the trial begins when the past piece is first placed into the mouth and ends when the last of the past is released from the pores and disappears into the mouth.
An adjustment is counted each time the pore releases and recontacts the past piece. When there's reformation of the pore hold or when there's an extension flexion or abduction, abduction of the digits, each adjustment is recorded separately for the left and right pore. Adjustments are not counted when the pore slides across the masterpiece with no movement of the digits or repositioning of the handhold.
Movements of the poor when it is not in contact with the pastor piece are also not counted. The time to eat is recorded for each trial starting with onset and ending when the pastor disappears into the mouth. If the animal pauses eating but maintains a hold on the piece, the time pause is subtracted from the total time the trial is omitted from the analysis.
If during any part of the eating, the view of the pause is obscured or if the animal breaks the piece, because shorter pieces alter handling patterns, mice sometimes drop the pasta during eating and after a short break, pick it up and resume eating. This may be due to distraction or partial satiety. If eating resumes quickly, we do not omit these trials, but subtract the time that pieces drop from the total eating time.
The measurements obtained from video analysis are the average number of adjustments per pore, per piece, the average eating time per piece, and the rate of adjustments. Now let's take a look at some atypical handling patterns. Atypical handling patterns can distinguish between different CNS injury types and reveal age related changes.
And recording these behaviors is useful as a supplemental analysis. Intact animals typically initiate eating with one poor place lower on the pasta and the other poor place near the mouth. The POS typically assume the same positions in all trials.
In an atypical behavior, the animal may switch positions of the two limbs. In the midst of eating in intact animals, the pores move into more symmetrical positions side by side or with digits interposed when the piece is very short. Some animals with CNS injuries will fail to do so, and rather will place the poles together in a symmetrical hold when the past pieces long.
Two additional atypical behaviors in rats and mice are failing to contact the pasta piece with one pore for an extended time during eating and using the mouth to pull the pasta through the pores to reposition them. Some behaviors are considered atypical for rats, but atypical for mice like hunched posture and dropping the pasta after eating begins. However, the frequency of drops in mice may increase after CNS damage, so it is useful to record drop frequency in both species.
Aged mice often make adjustments that do not result in advancement of the pasta. This is counted each time an adjustment clearly fails to result in a discernible movement of the pasta into the mouth. Now we'll show some results from the handling tests.
Analyses from rats with sensory motor cortical lesions indicate that animals make fewer adjustments with the impaired contralateral to lesion pull than with the ipsilateral po. Unilateral middle cerebral artery occlusion results in both a decrease in adjustments with contralateral limb and an increase in atypical handling behaviors. Video analyses from mice show that aged animals take longer to eat the pasta pieces and make more adjustments that do not result in advancement of the pasta into the mouth compared to young adult mice.
We've just showed you how to quantify pasta handling behaviors in rats and mice. When doing this procedure, it's important to remember to check interrater reliability to be sure that pasta adjustments are being accurately identified. So that's it.
Thanks for watching and good luck with your experiments.
