A SUITE OF WEB TOOLS FOR A SUITE OF WEB TOOLS FOR MORPHOMETRIC, GENETIC, AND MORPHOMETRIC, GENETIC, AND

MICROARRAY MICROARRAY ANALYSIS OF THE MOUSE CNSANALYSIS OF THE MOUSE CNS

Glenn Rosen, Melburn Park, Robert Glenn Rosen, Melburn Park, Robert Williams, Jonathan Nissanov, and Ken Williams, Jonathan Nissanov, and Ken

ManlyManly

Informatics Center for Mouse Informatics Center for Mouse NeurogeneticsNeurogenetics

• The Mouse Brain LibraryThe Mouse Brain Library - - RosenRosen

• iScopeiScope -Williams, Park -Williams, Park• NeurocartographerNeurocartographer - Nissanov - Nissanov• Neurogenetics Tool BoxNeurogenetics Tool Box - -

ManlyManly

Goal of ProjectGoal of Project

• Provide online resource for Provide online resource for complete mapping of complete mapping of differences in brain structure differences in brain structure to defined sets of genes to defined sets of genes (quantitative train loci; QTLs)(quantitative train loci; QTLs)

• Large scale (e.g., brain weight, Large scale (e.g., brain weight, volume)volume)

• Small scale (e.g., neuron number)Small scale (e.g., neuron number)

Specfic ImplementationSpecfic Implementation

• Universal Access to Universal Access to Neuroanatomic PhenotypesNeuroanatomic Phenotypes

• Digital DissectionDigital Dissection• Comprehensive Genotype Data Comprehensive Genotype Data

and Tools for Complex Trait and Tools for Complex Trait AnalysisAnalysis

Universal Access to Universal Access to Neuroanatomic PhenotypesNeuroanatomic Phenotypes

• The Mouse Brain LibraryThe Mouse Brain Library• Digital images (0.1–25 µm/pixel) from Digital images (0.1–25 µm/pixel) from

inbred, RI, and advanced intercross inbred, RI, and advanced intercross mice.mice.

• Ideal for stereologic analysis.Ideal for stereologic analysis.

• iScopeiScope• High-power, through-focus movies.High-power, through-focus movies.• Microscope controlled over the internet.Microscope controlled over the internet.• Ability to perform neuronal counts on Ability to perform neuronal counts on

selected regions of selected sections.selected regions of selected sections.

Digital DissectionDigital Dissection

• NeurocartographerNeurocartographer• Generate 3D atlas of mouse brain.Generate 3D atlas of mouse brain.• Use 3D atlas to automatically Use 3D atlas to automatically

segment regions of brain.segment regions of brain.

Comprehensive Genotype DataComprehensive Genotype Data

• DatabasesDatabases• RI genotypes and phenotypesRI genotypes and phenotypes• Advanced intercross progeny (GAdvanced intercross progeny (G1010))• Microarray data from inbred and RI Microarray data from inbred and RI

strainsstrains

• Neurogenetic Tool BoxNeurogenetic Tool Box• QTLs without tissue or DNA (WebQTL)QTLs without tissue or DNA (WebQTL)• Statistically evaluate neuroanatomic Statistically evaluate neuroanatomic

and microarray data with these DBsand microarray data with these DBs

The Mouse Brain The Mouse Brain LibraryLibrary

Gene Discovery by QTL Gene Discovery by QTL MappingMapping

•Quantitative Trait LociQuantitative Trait Loci

• Many medically interesting traits are complex Many medically interesting traits are complex traitstraits

• Complex = multiple genes and non-genetic Complex = multiple genes and non-genetic factorsfactors

• Genes which control complex traits are QTLsGenes which control complex traits are QTLs

•QTLs are detected by statistical associationsQTLs are detected by statistical associations

• Trait values Trait values marker locus genotypes (or marker locus genotypes (or estimated genotypes between markers)estimated genotypes between markers)

• Potential QTL indicated by LOD score maximumPotential QTL indicated by LOD score maximum

• Significance measured by LOD score valueSignificance measured by LOD score value

QTL Mapping PrinciplesQTL Mapping Principles

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TraitMarker genotype

Pro

geny

indi

vidu

als

Evaluate association for many markers and locations between markers

map location

sign

ific

ance

QTL Mapping GoalsQTL Mapping Goals

• SignificanceSignificance• Which LOD score maxima Which LOD score maxima

correspond to statistically correspond to statistically significant QTLs?significant QTLs?

• LocationLocation• How precisely does the LOD How precisely does the LOD

score maximum define the QTL score maximum define the QTL location? location?

• ArchitectureArchitecture• How many QTLs and what How many QTLs and what

interactions determine a interactions determine a particular trait?particular trait?

Permutation Test for Permutation Test for SignificanceSignificance

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TraitTraitMarker Marker

genotypegenotype

P = 0.05

LRS statistics from 1000 LRS statistics from 1000 regressions with permuted regressions with permuted

datadata

Bootstrap for LocationBootstrap for Location

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TraitMarker genotype

.

. 200 repetitions

.

Lateral VentriclesLateral Ventricles

Distribution of Ventricle SizeDistribution of Ventricle Size

0

1

2

3

4

Per

cent

Ven

tric

le

A/J

C57

BL /

6J

AX

B4

BX

A8

BX

A7

BX

A4

AX

B8

AX

B6

BX

A17

AX

B15

AX

B5

BX

A16

BX

A14

AX

B13

AX

B20

AX

B12

BX

A2

BX

A12

BX

A1

BX

A13

BX

A11

BX

A24

BX

A25

AX

B10

AX

B2

BX

A26

AX

B1

AX

B24

Strain

Mapping Ventricle SizeMapping Ventricle Size

0

5

10

15

20

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 X

LR

S S

core

LR

S S

core

ChromosomeChromosome

Epistatic InteractionsEpistatic Interactions

The iScopeThe iScope

Two Goals for the Online Two Goals for the Online MicroscopeMicroscope

• Web publication of Web publication of hardhard slide slide librarieslibraries• MBLMBL• Pathfinding for other collectionsPathfinding for other collections

• Automatic production of Automatic production of softsoft librarieslibraries

Java InterfaceJava Interface

• Navigation Navigation controlscontrols

• High and low High and low power index power index viewsviews

• Quicktime Quicktime screenscreen

Control PaneControl Pane

Navigation PanesNavigation Panes

• Blue crosshairs Blue crosshairs show current show current locationlocation

• Moveable red Moveable red dot selects dot selects desired locationdesired location

SchedulingScheduling

Co-development: Focussing Co-development: Focussing ProblemProblem

• Automatic focussing via Automatic focussing via proprietary noise-analysis proprietary noise-analysis algorithmalgorithm

• Interaction with vendor to Interaction with vendor to obtain custom chip giving us obtain custom chip giving us software access to noise valuesoftware access to noise value

• Focussing can be taken on- of Focussing can be taken on- of off-board off-board

Robotic Slide LibraryRobotic Slide Library

• Commercial Commercial designdesign

• Smartmotor™ Smartmotor™ technologytechnology

• Pneumatic Pneumatic actuatorsactuators

• Changeable Changeable cassettescassettes

• 6x82 2x3 inch 6x82 2x3 inch slidesslides

NeurocartographerNeurocartographer

The Navigation ProblemThe Navigation Problem

TT

T

T

T

Atlas (C57BL/6J Male)Atlas (C57BL/6J Male)

• www.neuroterrain.orgwww.neuroterrain.org• www.www.nervenetnervenet.org.org

Atlas AvailabilityAtlas Availability

2D-2D Rigid-body Alignment

BlockfaceImages

SectionImages

Median Filter

AIR

Error Detection

Reinitiali-zation

AIR

Acceptable? Resample

Reconstruction

SectionImages

Alignment Parameters

Y

N

MacroVoxel ArchitectureMacroVoxel Architecture

NetworkConnectionNetworkSlicingEngine

AtlasViewNavigation

ControlVOI Display

ManagementMouse

TrackingMovement controlsSlicing Axis controlTrack resultsReturned imageReslicecommandsVOI displayparameters

Neurocartographer ClientNetworkConnectionSlicingEngineAtlasData

Model

K2 CommandRedirection

ClientParameterSupport

Reslice commandClient displaysettings

K2 controlsclient displayNeurocartographer ServerK2 softwareNetworkConnection

DatabaseConnectionFederationImage data accessMetadata

access

Server-Client MacOStatServer-Client MacOStat

Ordered Unaligned 2D 2.5D

Auto Manual

Matching Plane Pairs

Matching Inplane Features

3D Reconstruction

Aligned Sections

Auto Manual

Z-A

xis

mat

chin

gX

Y P

lane

mat

chin

gW

arpi

ng

Matching 3D Features

3D

ParcellationParcellation

WarpingWarping

refere

nce

test

test

Case 1:

Case 2:

ResultsResultsCase 1 Case 2

Next StepNext Step

Neurogenetics Tool Box Neurogenetics Tool Box (WebQTL)(WebQTL)

WebQTLWebQTL

Trait data from MouseBrain Library images

UsersTrait data from

Neurocarto-grapher

User traitdata

Trait data from MouseBrain Library images

UsersTrait data from

Neurocarto-grapher

User traitdata

WebQTL Mapping ServiceWebQTL Mapping Service• http://http://

webqtl.roswellpark.org/webqtl.roswellpark.org/• Mapping populationsMapping populations

• Recombinant inbred linesRecombinant inbred lines• BxD advanced intercrossBxD advanced intercross

• No genotyping necessaryNo genotyping necessary

WebQTL Data EntryWebQTL Data Entry

• Choice of data entry Choice of data entry methodsmethods• Choose a data fileChoose a data file• Paste or type dataPaste or type data

WebQTL Data ConfirmationWebQTL Data Confirmation• Data entry or Data entry or

confirmation formconfirmation form

Mapping ProceduresMapping Procedures

Interval MappingInterval Mapping

Interval Mapping DetailInterval Mapping Detail

Future DirectionsFuture Directions

• MBLMBL• Extend MBLExtend MBL• Improve DB integrationImprove DB integration• Interface enhancementInterface enhancement• On-line image analysis On-line image analysis

toolstools

• iScopeiScope• Additional design and Additional design and

constructionconstruction• Robotic slide handlingRobotic slide handling• Web archive of through-Web archive of through-

focus image stacksfocus image stacks• Streaming VideoStreaming Video

• NeurocartographerNeurocartographer• Strengthen and extend Strengthen and extend

segmentation algorithmsegmentation algorithm• Segment MBLSegment MBL

• Neurogenetics Tool Neurogenetics Tool BoxBox• Batch submission of Batch submission of

multiple traitsmultiple traits• Search for interacting Search for interacting

QTLsQTLs• Data archiving and exportData archiving and export• Additional crosses and Additional crosses and

cross designscross designs• DNA ArrayDNA Array

AcknowledgmentsAcknowledgments

• This project is part of the Human Brain Project/Neuroinformatics program funded jointly by the National Institute of Mental Health, National Institute on Drug Abuse, and the National Science Foundation (P20-MH 62009).

MBL Intro

MBL BD

Mbl Small

MBL All

Slide

MBL High

Power