.. raw:: html
.. _reproduction:
Experiment replication
======================
One of the main strengths of Stytra is the possibility of
sharing the experimental paradigms described in a publication
as scripts that can be run on different platforms and experimental
hardware. To prove the validity of this approach, we decided to showcase
the software reproducing the results from two publications that investigated
different behaviors of the larval zebrafish. This allowed us to
verify the performance of our package in producing and monitoring
reliable behavioral responses, and showed how the Stytra platform
can be used to share the code underlying an experimental paradigm.
The scripts used for designing these experiments are available in
our repository, together with a full list of parts and description
of the hardware. In this way, everyone can independently replicate
the experiments simply by installing and running Stytra on a suitable
behavioral setup.
Closed-loop motor adaptation
----------------------------
To demonstrate the effectiveness of the closed-loop stimulation
software for head-restrained larvae, we re-implemented in Stytra
one of the paradigms described in :cite:`portugues2011adaptive`.
This paper addresses the importance of instantaneous visual feedback
in the control of the optomotor response in seven dpf zebrafish larvae.
In :cite:`portugues2011adaptive`, a closed-loop paradigm was used to
have real-time control over the visual feedback that the animal receives
upon swimming. After triggering motor activity with forward-moving black
and white gratings (10 mm/s, 0.1 cycles/mm), online tail tracking was used
to estimate the expected velocity of the fish based on freely-moving
observations, and a backward velocity proportional to the expected
forward velocity was imposed over the forward grating speed. In one
crucial experiment (Fig 3 of :cite:`portugues2011adaptive`) the authors
demonstrated that reducing or increasing the magnitude of this velocity
by a factor of 1.5 (high gain) or 0.5 (low gain) resulted in modifications
of the bout parameters such as bout length and inter-bout interval
(time between two consecutive bouts). The figure below
shows the inter-bout interval along the protocol,
where the three gain conditions were presented in a sequence
that tested all possible gain transitions. When the gain increased
the fish was consistently swimming less (higher inter-bout interval),
while the opposite was observed when the gain decreased. Therefore,
as expected, fish adapted the swimming parameters to compensate for
changes in visual feedback. The line represents the average normalized inter-bout time, and bars represent standard error of the mean from n=28 larvae. Here is the figure adapted fron adapted from :cite:`portugues2011adaptive`).
.. figure:: ../../figures/portugues2011.png
:scale: 50%
:align: center
We reproduced exactly the same protocol within Stytra, and we used
Stytra modules for closed-loop control of a visual stimulus to
compare whether it could replicate the findings from
:cite:`portugues2011adaptive`. The cumulative angle of the
extracted tail segments was used with a gain factor to estimate
the fish velocity and the gain factor was changed in a sequence matching
the protocol in :cite:`portugues2011adaptive`. The replication with Stytra
yielded the same result:
inter-bout interval decreased in low gain conditions and increased in
high gain conditions. Here are plotted average and individual fish (n=24 larvae):
.. raw:: html
:file: ../../figures/portugues2011_replication.html
Closed-loop phototaxis assay
----------------------------
To test the freely swimming closed-loop performance,
we replicated a protocol from :cite:`huang2013spinal`. The fish is induced
to perform phototaxis by keeping half of its visual field (the left or the
right side) bright while the other is dark. The fish is more likely to
turn to the bright side. The stimulus is constantly updated so that the
light-dark boundary is always along the mid-line of the fish. We replicated
the qualitative trends observed in :cite:`huang2013spinal`, however the
ratios of forward swims to turns are notably different
(figure below). The variability of fish responses and
differences in the stimulus presentation setup (e.g. projector brightness)
could account for these differences. Also, to reduce duration of the
experiments, we included a radially-inward moving stimulus that brings
the fish back into the field of view.
.. image:: ../../figures/huang2013replication.png
Comparison of turning angle distribution in a closed-loop
freely-swimming phototaxis experiment.
Left: a histogram of the angle turned per bout, redrawn from
:cite:`huang2013spinal`.
Right: the equivalent panel, with n=10 fish and the protocol run with
Stytra. The dark shading on the plot represents the dark side of the visual
field.