as application

[as tRAVEL GRANT APpLICATION- christian jaques hissom] February 8, 2014

Investigating the functional significance of corticospinal and thalamocortical neurons in learning and performance of complex behavioral tasks in the adult

mammalian motor system

Abstract Contrary to popular perception, our brains are not simply static systems that process and respond to external stimuli, but are constantly adapting themselves in a dynamic fashion. One area of the brain that has been an ideal model for studying learning and associated brain plasticity is the primary motor cortex. In this study, our goal is to evaluate how distinct populations of neurons in the thalamus and primary motor cortex contribute to learning and performance of skilled motor tasks in rodents. Thalamocortical inputs to the motor cortex are postulated to relay information from a variety of nuclei that mediate and coordinate motor performance, including the basal ganglia and cerebellum. Corticospinal neurons are the sole source of neuronal output to the spinal cord from the cortex and are postulated to play a direct role in skilled motor learning in rats. Prior studies attempting to define the function of these neuronal systems in skilled motor behavior have involved ablation strategies that resulted in damage to non-target cell populations, thus confounding the interpretation of experimental findings. The goal of the present study is to use a novel viral approach to selectively eliminate distinct populations of neurons implicated in motor function and examine subsequent changes in motor performance. It is postulated that behavioral changes will occur that restrict proper task performance and inhibit motor adaptation.

Above is the condensed form of the project paper that was produced after completing the first round of experiments performed on subject rats during the summer portion of my research. I will also provide a copy of the introduction of the research paper which includes, in my words, an explanation of why we selected to perform a thalamocortical ablation.

I will briefly describe the several steps and procedures performed during the development and analysis of this project. These steps, outlined, are as follows: train rats in forelimb reaching task, inject toxin, repeat forelimb task, perform further behavioral assays, perfuse rats, remove brain and slice with microtome, perform immunohistochemistry, analyze ablation, collect and analyze data.

As a MARC scholar I was granted the opportunity to be an intern in the Tuszynski lab, UCSD Center for Neural Repair, under the supervision of Dr. James Conner, associate project scientist . My research experience began with a brief overview of the project and a journal filled folder. I was instructed to read and analyze the papers and to speculate the outcome of the project. The first two weeks of the project consisted of training, familiarization with the protocol and required techniques, and reading roughly 30 scientific journals. My room decorated with the contents of that folder and my whiteboard representing the intricate connections of the brain and culminated propositions; I was determined to have the correct answer.

I first learned how to use a microtome. This tool is used to slice 40 micrometer brain sections that are later mounted on a stage. Spare brains were used for training and soon enough I was confident in my ability to use this machine. Next, the slices were collected and stained using immunohistochemistry. During this process, the tissue is stained with antibodies that, once attached to a tagged tracer, expose the effects of the toxin used to ablate the desired brain cells.

After mastering the complexity of tissue processing I progressed to learn rat training. Tis required completion of VA medical hospital required training and paperwork; a process that takes up to three weeks. During this training session I learned how to care for, acclimate, and train the rat subjects used in almost every lab.


Every day, for about 8 hours a day, I would meet with Dr. Conner and assist with training. After training, the rats were surgically injected with the toxin. I was in charge of re-training the rats, post-surgery, in the forelimb reaching task. A series of behavioral assays were also performed in order to obtain a better understanding akin to the neurological difficulties observed.

The data was recorder, graphed, and analyzed. Furthermore, video footage was obtained in order to analyze each movement, frame by frame, with an attempt to pinpoint the potential nuclei that was malfunctioning. Hours were spent discussing potential candidates and formulating ideas and explanations that would then be justified by tissue processing.

Once training and data collection was finalized, the animals were perfused and tissue processing began. Eighteen rats were processed and we were able to see the outcome of the ablation roughly 2 months after the project began. This information was gathered and synthesized into an abstract and final project paper.

Over summer I worked 40 hours a week to assist with this great research project. I currently still work with Dr. Conner and the project has moved forward to analyze different connections akin the network of neurons that partake in the forelimb reaching task. The lab has adopted new techniques that allow researcher to inhibit or excite a specific population of target neurons by use of a designer ligand that works on a novel receptor. The use of this technique has just recently been introduced and I look forward to observing the results and performing behavioral assays. In this manner we can observe behavioral deficits that arise from both the loss and the strengthening of the thalamacortical tract. Aside from the thalamocortical and corticospinal tract, we will continue to examine connections within the network that correlates to the forelimb reaching task. Our next goal will be to analyze the connection that channels coordination and motor control from the cerebellum to the thalamus by performing a dentate-thalamic ablation.


Logistics

Purpose: To present and explain the procedure and outcome of research conducted under the supervision of Dr. James Conner. This opportunity will strengthen my ability as a scientist to create and organize a presentation and to communicate the complexity of this project in a clear and consist manner.

Conference: Emerging Researchers National Conference in STEM February 20-22, 2014 Location: Washington D.C Contact: Donna Behar, ERN coordinator - [email protected] Conference description: The ERN conference is designed to promote scientific enrichment.

Topics in STEM converge to create an experience that, I believe, will strengthen and enrich my scientific career. Conference events include: Plenary session, guest speakers, poster presentations, oral presentations, exhibitions, graduate student presentations, and networking events. Budget

Expenses Expense Detail Expense Detail TotalFlight - I will leave on Thursday night the 20th

and return Sunday afternoon, the 23rdAirfare quoted amount: $557.91

USDTaxes and fees:$85.84 USD

$643.75 USD

Conference registration fee -Confirmation Number: M7NZDY7NJ56 $400 USD

Lodging -Four Points by Shareton -$107/night + Taxes-Friday + Saturday night $245.49 USD

Transportation -Taxi: UCSD to San Diego- Lindbergh Intl Arpt (SAN)

-Taxi: Lindbergh Intl Arpt (SAN) to UCSD to San Diego

-Taxi: USD Ronald Reagan National Arpt (DCA) to Hotel (1201 K St NW Washington,

DC 20005)

-Taxi: USD Hotel to airport

-Bus transportation:From hotel to conference

– $49.6515 miles

– $49.6515 miles

– $24.174.9 miles

-$24.174.9 miles

-$2.42 USD/ trip$9.68 USD expected

$147.64 USD

Food -Saturday- Sunday Breakfast, Lunch and dinner

-Washington D.C average cost/meal $15 USD ( $11-$20

USD)

$120 USD

Poster Printing -$40 USD $40 USDTotal $ 1436.88 USD

mailto:[email protected]


IntroductionNeuroplasticity is the antagonist of the once held dogma; that the brain, once fully developed, remains stagnant in its ways and resilient to change. It is know now, however, that behavioral changes and learning elicit new synaptic connections that rearrange neural pathways. This phenomenon occurs throughout the entirety of the brain which makes it difficult to study. However, we can isolate this phenomenon by observing changes that occur in the brain due to motor learning.

The basal ganglia and cerebellum are two important brain regions necessary for learning and motor performance. These brain regions do not communicate directly with primary motor cortex but relay information to the primary motor cortex (M1) by way of the ventral lateral and ventral anterior (Va/Vl)nuclei of thalamus (Cruikshank et al 2010). The basal ganglia has been postulated to play a role in motivation, motor control, gain control, and motor learning. The cerebellum corrects and plans motor commands and coordinates proper balance (Kaneko et al 2009). Output from these distinct brain centers converges onto Va/Vl and is then relayed to primary motor cortex where thalamocortical inputs terminate monosynapticly onto corticospinal motor neurons in Layer VB.

While prior studies ascertain the general function of these brain regions, the complexity and significance of each individual connection that collaborates in the overall circuit of each brain region, be it cerebellum or basal ganglia, remains incomplete.

Prior studies rely on electrolytic or cytotoxic lesions that not only damage the target but also elicit collateral damage to surrounding areas Thus, a significant problem which stems from such lesions—difficult as it may be to target a specific area in the brain, it is even more difficult to target a specific connection and analyze a circuit in the brain.

The present experiment makes use of a novel method that allows for highly selective ablations of a specific connection in the brain. A rabies pseudotyped lentivirus is used to retrogradely infect cells providing afferent innervation of a specific brain region with a foreign receptor, the human form of the IL2 receptor. Later, an immonotoxin selectively targeting the human IL2 receptor is used to mediate selective ablation of one or more of the infected cell populations.

Using this technique we can hypothesize and observe the functional significance of the thalamocortical tract akin to a complex motor behavior such as distal forelimb reach and dexterity. In this manner, learning and performance related brain regions can be disassembled and analyzed in order to ascertain the origins of motor learning and task performance.

Hypothesis: It is postulated that a thalamocortical ablation should result in behavioral irregularities parallel to neurological disorders such as Parkinson’s and Huntington’s disease ataxia, tremors, and dysdiadochokinesis, respectively. Therefore an overall decline in forelimb reaching performance, inability to adapt to task altercations, and uncoordinated movements are expected.


Trip Overview

Trip Name: Trip from San Diego to Washington (For Christian Jaques Hissom )Start Date: Feb 20, 2014End Date: Feb 23, 2014Created: Feb 3, 2014, KIRSTEN KUNG (Modified: Feb 3, 2014)Description: ERN Conference -- HissomPlease enter 00 and your 6 digit Event Number, the TEPC number, or Personal travel enter 00999999.: 00902655Purpose of trip: BusinessAgency Record Locator: KFGSBFPassengers: Christian HissomTotal Estimated Cost: $643.75 USDAirfare must be ticketed by an agent by: 02/04/2014 11:00 PM Pacific

Balboa Travel (University of California - San Diego campus) 800-682-6170800-359-8773

Reservations

Thursday, February 20, 2014

Flight San Diego, CA (SAN) to Atlanta, GA (ATL)Delta 1792This flight leaves on Feb 20 and arrives on Feb 21.

Departure: 10:50 PMSeat:25E (Confirmed)Lindbergh Intl Arpt (SAN)Terminal:2Duration: 3 hours, 54 minutesNonstopArrival: 05:44 AMHartsfield Intl Arpt (ATL)Terminal:SOUTH TERMINAL

Confirmation: HCYHRTStatus: Confirmed

Additional Details

Aircraft: Boeing 737-900 Distance: 1885 miles

E-Ticket

Cabin: Economy (U) Meal: Refreshments for Purchase

Friday, February 21, 2014

1 hr, 36 min layover at Hartsfield Intl Arpt (ATL)Flight Atlanta, GA (ATL) to Washington, DC (DCA)Delta 1138Departure: 07:20 AMSeat:34A (Confirmed)



Hartsfield Intl Arpt (ATL)Terminal:SOUTH TERMINALDuration: 1 hour, 39 minutesNonstopArrival: 08:59 AMRonald Reagan National Arpt (DCA)Terminal:B

Additional Details

Aircraft: Douglas MD-85 Distance: 547 miles

E-Ticket

Cabin: Economy (U)

Sunday, February 23, 2014

Flight Washington, DC (DCA) to Minneapolis/St. Paul, MN (MSP)Delta 1763Departure: 02:35 PMSeat:22D (Confirmed)Ronald Reagan National Arpt (DCA)Terminal:BDuration: 2 hours, 48 minutesNonstopArrival: 04:23 PMMinneapolis St Paul Intl (MSP)Terminal:TERMINAL 1 - LINDBERGH


Additional Details

Aircraft: Douglas MD-90 Distance: 928 miles

E-Ticket

Cabin: Economy (X) Meal: Refreshments for Purchase

1 hr, 17 min layover at Minneapolis St Paul Intl (MSP)Flight Minneapolis/St. Paul, MN (MSP) to San Diego, CA (SAN)Delta 1389Departure: 05:40 PMSeat:37B (Confirmed)Minneapolis St Paul Intl (MSP)Terminal:TERMINAL 1 - LINDBERGHDuration: 3 hours, 51 minutesNonstopArrival: 07:31 PMLindbergh Intl Arpt (SAN)Terminal:2


Additional Details

Aircraft: Boeing 757-200 Distance: 1529 miles

E-Ticket


Cabin: Economy (X) Meal: Food for purchase

Total Estimated CostAir

Airfare quoted amount: $557.91 USD

Taxes and fees: $85.84 USD

Total Estimated Cost: $643.75 USDRestrictionsQuote: NONREF/PENALTY/APPLIESTICKET NOT YET ISSUED. AIRFARE QUOTED IN ITINERARY IS NOT GUARANTEED UNTIL TICKETS ARE ISSUED.

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