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Studying Learning and Memory in Biological

Neural Cell Networks In Vitro by Mimicking


In Vivo Conditions
Abhishek Kumar
December 9, 2010

1 Introduction
Learning and memory are the most remarkable features of brain. But still
we know very little about how brain actually achieves them. Major obstacles
in studying them are difficulty in recording in vivo, designing simple but
logical behavioral experiments and above all doing the experiments without
too much disturbing the normal routine of the animal under observation.
In recent past many people have been able to cross these hurdles to some
extent[5].
Assuming the above mentioned challenges are tackled, the extent of free-
dom in observing the flow of various signals and then to conclude any sub-
stantial remark about the processes involved in learning and memory while
doing in vivo experiments is very limited and unsatisfactory. Also the sys-
tem (animal’s brain) will undergo so many changes during the observation
that any repeated experiment followed by in vivo recordings will give highly
distorted inferences about the system’s response.
These challenges have crippled the advancements in understanding learn-
ing and memory of brain. There have been efforts to study simpler systems
involving less number of neurons but designing suitable behavioral experi-
ments for them is a challenge.
Proposal: To study learning and memory in biological neural cell net-
works in vitro by mimicking in vivo conditions.

1
In recent years, many people have successfully demonstrated fully in vivo
neural signal recording and stimulation systems[7, 9]. Such systems have been
very remarkable in retinal, cochlear and other prosthesis applications. They
can be used to create database of sensory inputs coming to brain during well
planned behavioral experiments by recording in vivo from strategic locations.
Neurons cultured from new born as well as adult animals have been found
to survive for one month or more in normal conditions[6, 2]. Database of
sensory inputs created from in vivo recordings can be used as input signals
to train cultured biological neural cell networks by suitably choosing few
neurons to receive inputs. Physiological environment will be provided to the
cultures to mimic in vivo conditions during training.
Top down approach is proposed to study the system at each level. Firstly,
the network and microcircuit involved in the learning and memory will be
studied. As there are methods available to track firing neurons[8], it is possi-
ble to trace the subset of neurons which are participating in learning and
memory of corresponding experiment. Then individual neurons showing
changes in firing pattern during learning will be carefully analyzed. These
analysis will be much more detailed due to in vitro experiment.

1.1 Key questions


1. Is it possible to train smaller set of neurons to perform any task of
complete brain?

2. How the network changes during different phases of learning?

3. How information is encoded spatially in the culture?

4. How different cultures of neurons learn and store memory for same
inputs?

5. Is it possible to train similar culture of neurons for inputs from different


sensory organs? If yes, then is there any resemblance in the architecture
of networks so obtained?

6. How neurons cultured from newly born animals differ from that of adult
animals during learning?

7. What kinds of plasticity are involved for different kinds of cultures?

2
It is expected that nature uses the most optimized method for solving any
problem. Therefore, the basic architecture behind learning and memory will
be same for neural networks inside brain or outside brain. So, we can analyze
actual system better after understanding the architecture. Also, knowledge
about architecture behind learning and memory can be exploited to make
intelligent machines.

2 Objectives
Long term objective of this proposal is to develop better understanding of
processes behind learning and memory in brain. Some of the short term goals
are given below:-

ˆ To create a database of in vivo sensory inputs recorded during well


planned behavioral experiments.
First target of the project will be visual system. Behavioral experiments
involving simple pattern recognition will be developed. A neural signal
recording chip with large number of recording sites will be implanted
near retinal cells[9]. A database of well documented experimental setup
(preferably video recording) and recorded signals will be created. Af-
ter the creation of database for visual system, it can be extended to
auditory system also.

ˆ To culture long lasting neurons and stimulate them with signals stored
in database.
Initially neurons will be cultured from the system corresponding to the
recorded signal. Special techniques will be employed during culture so
that firing neurons can be tracked during stimulation. Also the phys-
iological conditions will be maintained similar to that present inside
brain. Neurons with less inputs will be used as starting points of net-
work. Growth of network will be observed during training. Response
of the network will be checked for test inputs.

ˆ To investigate single neurons for any biological changes after training.


Population of neurons with similar structure containing neurons partic-
ipating in learning as well as isolated during stimulation will be inves-
tigated. Using standard protocols and methods they will be checked

3
for variety of changes[1, 4]. Some methods will need special kind of
cultures.

3 Work Plan
Early stage of the project will involve selecting suitable system. Brain of a cat
is a good system for visual experiments[10]. Suitable behavioral experiments
involving simple pattern recognition will be developed. Neural recording
chips will be implanted near retinal cells. Same experiment will be performed
on cats with different ages. All the information related to experimental setup
and recorded signals will be used to create a reusable and sharable database.
Size of the database will keep on increasing with time and it will be helpful
to many other labs also working in neurophysiology.
Next phase will consist of culturing neurons from diverse systems. Cul-
tures will be kept at body temperature and surrounding solution will be made
identical to what present in the brain. Input stage of the training network
will be formed by neurons without any input from other neurons. As a result
inputs to subsequent stages will be similar to what a cell receives inside brain.
Recorded signals will be given to input stage neurons through stimulation
electrodes.
Tracking of firing neurons during and after learning phase will give the
information about network architecture responsible for learning and mem-
ory. So, designing an efficient way to track neurons without disturbing them
is very crucial. Such an experiment has been done by replacing the coding
sequence of Arc/Arg3.1 gene with that of the gene that encodes green fluo-
rescent protein (GFP)[8]. A lesser disturbing approach is to culture neurons
on micro electrode array with small size electrodes so as to achieve better
resolution[3]. But this will limit the number of cells which can be tracked.
A mix of above two methods is proposed to track the firing neurons.
As the neurons are available in vitro, they can be analyzed for changes oc-
curring due to learning. Neurons which have not participated during training
or neurons from fresh culture will be used to set control value. Now cells with
similar structure but actively participating during training will be analyzed
for different kinds of plasticity occurred due to learning[1, 4].

4
References
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Functions, and Mechanisms. Neuropsychopharmacology, pages 18–41,
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[2] Lars Eide and Cynthia T Mcmurray. Culture of adult mouse neurons.
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[3] Y Ito, T Yagi, H Kanda, S Tanaka, M Watanabe, and Y Uchikawa. Cul-


tures of Neurons on Micro-electrode Array in Hybrid Retinal Implant.
Investigative Ophthalmology, pages 414–417.

[4] Sang Jeong Kim and David J Linden. Ubiquitous plasticity and memory
storage. Neuron, 56(4):582–92, November 2007.

[5] Jeffrey M. Perkel. Cell signaling: in vivo veritas, 2009.

[6] Steve M Potter and Thomas B Demarse. A new approach to neural cell
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[9] James D Weiland, Wentai Liu, and Mark S Humayun. Retinal Prosthe-
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[10] Torsten N Wiesel. THE POSTNATAL DEVELOPMENT OF THE VI-


SUAL CORTEX AND THE INFLUENCE OF ENVIRONMENT. Nobel
Lecture, (December), 1981.