Manual

From www.temporal-lobe.com

Contents 1 Introduction 2 Working with the diagram 2.1 System requirements 2.2 Opening the diagram 2.3 The diagram interface 3 The diagram area 3.1 Area boxes: overview 3.2 Three-dimensional axes 3.3 Area boxes: details 3.4 Toolbar 3.5 Result of search 3.6 Anatomical information 3.7 Searching for Connections 4 Summary and example 5 Quality assurance 6 Appendix 1: Abbreviations 7 Appendix 2: Problem solving

Introduction

(This page is currently being updated.)

The interactive connectome allows you to customize the display of all known connections of the rat hippocampal formation, the parahippocampal and the retrosplenial cortex in a way that is most relevant to your specific field of interest. It is built upon a database which contains all reported anatomical connections in these regions in the rat. The database may eventually become published online on www.temporal-lobe.com, but is currently offline.

Features of the connectome:

• The connectome incorporates in a single display all known anatomical literature to date of one species (i.e. the rat). In many previous reviews, data from different species were combined into “an average” description. This connectome offers an objective description of current anatomical data.
• A search was performed on publications reporting tract-tracing studies on RSC-HF-PHR connections in PubMed (www.PubMed.gov) and Embase (www.embase.com). The following inclusion criteria were used: 1) Tract-tracing studies or studies which report intracellular filling of single cells; 2) Studies which used healthy, genetically un-altered, untreated adult rats were included; 3) Publications written or translated into English or in a language using roman print.
• The connectional information was retrieved from the papers, including information from tables and figures, using the following criteria: 1) It was clear where anterogradely filled terminals or retrogradely labeled cell bodies were located; 2) The location of the injection site was clearly described; 3) Injection sites did not include multiple brain areas or fiber bundles; 4) Lesion studies were discarded; 5) Explicitly reported non-excitatory projections were excluded; 6) Contralateral projections were excluded.
• All connections reported in the literature are included whereas in most reviews only the prominent connections are reported. In this connectome almost 2600 connections are present.
• The connectome is interactive; users can switch on and off connections between cortical layers in the network. This will allow users to better comprehend the impact of a lesion, make anatomically correct theoretical models of this region, and help interpret electrophysiological data of this common laboratory animal.
• The connectome points out gaps in anatomical knowledge. In the connectome, several levels of detail are available in order to be able to draw all reported anatomical connections. This enables users to quickly spot which connections are described in the literature with or without much detail.

Working with the diagram

System requirements

In order to open the pdf-file, you need to have Adobe Acrobat Reader installed on your system. Adobe Acrobat Reader is available for most operating systems. You can download Adobe Acrobat Reader free of charge by clicking on the image below:

It is recommended that you update your Acrobat Reader to the latest version. As of Acrobat Reader version 8, hardware accelerated drawing is introduced in the software which will speed up rendering of the diagram. Acrobat Reader prior to version 6 does not support layers, therefore it cannot display the diagram properly. When opening version 2 of the diagram, a warning message will be displayed if you are using an Acrobat Reader prior to version 9.

Opening the diagram

The diagram (right click -> save as works best) can be opened by Adobe Acrobat Reader. Please refer to the "System requirements" for obtaining a copy of Adobe Acrobat.

The diagram interface

After opening the connectome in Acrobat Reader, a view similar to that in Figure 1 should be visible on your computer screen.

The diagram area

Area boxes: overview

The connectome consists of 14 large, color-coded area boxes, which represent the sub-regions of the hippocampal formation, parahippocampal region and retrosplenial cortex. The area names are abbreviated in the connectome. The areas included are (figure 1A, white outline):

Hippocampal formation:

• Dentate gyrus (DG)
• CA3
• CA1
• Subiculum (Sub)

Parahippocampal region:

• Presubiculum (PrS)
• Parasubiculum (PaS)
• Lateral Entorhinal Area (LEA)
• Medial Entorhinal Area (MEA)
• Perirhinal cortex Area 35 (A35)
• Perirhinal cortex Area 36 (A36)
• Postrhinal cortex (POR)

Retrosplenial cortex

• Retrosplenial cortex Area 29a (A29a)
• Retrosplenial cortex Area 29b (A29b)
• Retrosplenial cortex Area 29ab (A29ab)
• Retrosplenial cortex Area 29c (A29c)
• Retrosplenial cortex Area 30 (A30)

Three-dimensional axes

The 14 anatomical sub-regions are three-dimensionally organized. However, the origin and termination of connections are not always described in full detail in the literature. Therefore, area boxes are divided into four quadrants (white dotted lines in Figure 1). Quadrant I has full topological information, whereas the other quadrants have less topological detail. In quadrant I, the vertical axis in the connectome represents the septotemporal axis of dentate gyrus (DG), Cornu Ammonis (CA3 and CA1), subiculum (Sub), presubiculum (PrS) and parasubiculum (PaS), and the dorsoventral axis of medial and lateral entorhinal area (MEA and LEA), retrosplenial cortex (A29 and A30), perirhinal cortex (A35 and A36) and postrhinal cortex (POR). The sidebars (Figure 1B, red boxes) display the dorsoventral and septotemporal axes of the anatomical sub-regions. The horizontal axis within quadrant I and III represents the proximodistal axis in CA3, CA1, Sub, PrS and PaS; the rostrocaudal axis in A29c, A30, A35, A36 and POR and the DG is subdivided into the inner/outer blades and crest region. In quadrant II, the information of the vertical axis and the layers are specified, but no details of the horizontal axis are presented. In quadrant III the horizontal axis and the layers are represented, while in quadrant IV, only layer information is present. Within the area boxes, the layers for each specific subarea are outlined.

Area boxes: details

In this paragraph, we will introduce the elements within the area boxes and their three-dimensional axes. The Sub and A35 will serve as examples.

Let us first take a look at the top part of the Subiculum area box (Figure 3). At the very top the word "Sub" is written, which is the abbreviation for Subiculum (See Appendix 1). In the next row, the words "prox", "int", "dist" and "unsp" are written. These labels specify the proximo-distal axis of the Sub, with "prox" meaning proximal, "int" meaning intermediate and "dist" meaning distal with respect to the DG. The label "unsp" (unspecified) indicates that no information is available on the proximo-distal distribution of these connections. Below the proximo-distal axis three rows of smaller boxes are displayed. These columns of smaller boxes represent the cortical layers that are defined in the Sub at the three levels of the proximo-distal axis or at the unspecified proximo-distal level. The layers in the Sub are the stratum moleculare and the stratum pyramidale. In the Sub the stratum moleculare is subdivided into a superficial (sup) and deep part of the layer.

Figure 4: Subiculum area box with septo-temporal and proximo-distal axis

As was explained in paragraph 2.4.1a, in addition to a proximo-distal axis, a septo-temporal axis is also present in the connectome (grey bar). Therefore, the cortical layers are repeated for the septal (top rows), intermediate (middle rows) and temporal level (bottom rows) of the Sub (Figure 4).

This information is once more repeated for the unspecified level of the Sub, in which no septo-temporal subdivision is made (Figure 5):

Figure 5: Unspecified level of the Subiculum area box

Toolbar

The interactive connectome allows visualization of detailed connectivity patterns within and between sub-regions. To search for connections, use the search button on the toolbar (Figure 1C – orange outline). The toolbar has 8 buttons (from left to right): Search for connections, show all connections, clear all connections, a short tutorial on how to search for connections, a help section, contact information, a link to the project website www.temporal-lobe.com and information about how to support this project. If the toolbar is closed, clicking the ‘open menu’ button will restore it (Figure 1D, green outline).

Result of search

After a search is carried out, the retrieved connections will be drawn in the connectome between the appropriate areas and quadrants. Additionally, an eye-icon will appear in the layers panel on the left (Figure 1E, yellow outline). This is an alphabetically sorted list of ‘from–>to’ connection groups that can also be switched on or off manually. In front of each group is a ‘+’ icon. Clicking this icon expands the list of individual connections that make up the group, allowing one to select connections originating from a specific cortical layer or according to a specific three-dimensional projection pattern.

Anatomical information

The figure panel (Figure 1F, blue outline) provides a detailed anatomical description of the retrosplenial cortex, the hippocampal formation and the parahipppcampal region, together with translation tables for nomenclature. Use the numbered buttons in the bottom right of this panel to switch between the figures.

Searching for Connections

There are two ways of showing connections in the connectome:

1. Use the search box, placed in the middle of the pdf;
2. Use the layer navigation pane.

* Search Method 1: Search box.

1. Open the search box
2. Select a brain area from the ‘Area’ listbox (top box in search window). For example, the lateral entorhinal area (LEA).
3. The search and display of the connections can be limited to a particular 3D subregion. For example, in the entorhinal cortex, a dorsolateral-to-ventromedial axis is described. In the 3D axis fields, a selection can be made for the available subdivision of the selected area. In the ‘3D axis 1’, you can select a ‘vertical axis’, e.g. temporal-to-septal, dorsal-to-ventral or dorsolateral-to-ventromedial axis. In the ‘3D axis 2’, you can select the ‘horizontal’ axis, e.g. the proximo-distal axis.
4. The search can also be limited in to include only a particular cortical layer. When you leave the ‘-‘ in the field, connections to / from all layers will be searched for.
5. Select ‘afferent’ of ‘efferent’. If you select ‘efferent, you get the connections that are running from the area selected in the first Area box. If you select ‘afferent’, you get the connections that are running to the area you selected in the first Area box. If you are interested in both the afferent and the efferent connections, you need to perform the search twice.
6. Search results can be filtered to include only a particular area.

If you want the add connections to your search, repeat the steps 1-6. If you want to start again, please first clear all connection (‘Clear all’ button in the toolbar).

* Search Method 2: Layer Navigation Pane

On the left hand side, the layers navigation pane should be visible. If no layers are visible, please check your version of Acrobat Reader (see System Requirements). The layers are alphabetically organized. At the top of the list, a checkbox ‘options’ is available. This layer must be checked, to see the connectome. to select or deselect the figures in the left top corner of the connectome.

In order to show or hide connections between areas, you need to use the layers navigation pane on the left side of the screen (Figure 1). By checking a box, you can switch on or off all the projections from a specific sub-area to another (Figure 6). The "eye"-icon appears when the connection is displayed in the diagram area. Layers that are marked with a "lock"-sign are inactive (for instance "areas" in Figure 6) and cannot be selected or excluded ("deselected").

By clicking the "+" icon in front for the projection of interest, the list of layer-by-layer connections expands and the "+" icon changes into a "-" icon. Clicking the "-" icon again will collapse all layers again under their main folder.

When the layer-by-layer connections are expanded (Figure 7), a list allows you to individually switch on and off connections. In order for Adobe to draw individual connections, the top checkbox (Figure 8), which in this example is Sub->A35, needs to be selected. This may mean that you need to deselect many individual layers when you wish to view only a specific layer in a "From->To" connection.

Supplement 2 (see References) provides a reference table in which the references between two specific areas can be looked up. In addition, a complete list of all used references with hyperlinks to "PubMed" is provided.

If you deselect the top checkbox, all individual layers underneath the deselected connection will be hidden from the diagram.

The layer names in the layer panel follow a regular syntax which explains what is drawn:

From -> To

Area_axis1_axis2_layer -> Area_axis1_axis2_layer

Area_axis1_layer -> Area_axis1_layer (only for LEA and MEA)

• Area: abbreviation of the area name
• Axis 1: septo-temporal axis, dorsolateral-ventromedial or dorso-ventral axis
• Axis 2: proximo-distal axis or rostro-caudal axis
• Layer: abbreviation of the cortical layer name

Summary and example

Figure 9 provides a summary of all the information that was presented step-by-step in Section 2. As an example the connections between Sub and A35 are shown and here we will give a short interpretation.

Selecting the connections: When selecting the A35 -> Sub check box in the layers pane, the connections from Area 35 to the Subiculum will appear. Likewise, when selecting the Sub -> A35 check box, the connections from Sub to A35 will appear. The details of these connections can be made visible in greater detail by looking at the layer to layer connections (see 3.4 and Figure 7). From this the following can be learned:

Sub->A35: from the diagram it follows that both septal and temporal levels of the Sub project to A35. However, not much detail of this connection is provided in the literature. Therefore, the origin of the connection in the Sub is drawn at the "unsp"-box at both septal and temporal levels. The connection arrives in the unspecified level of A35 because nothing has been reported about the dorso-ventral and rostro-caudal level of the termination. In fact, so little is known that the connection arrives in "unsp", which means that no information has been reported about the cortical layer in which this connection terminates. From Supplement 2 (see section 4: Quality assurance) it can be learned that this connection was reported in two references 2-3.

A35->Sub: From the layer-to-layer details it follows that the dorsal part of A35 projects to the deep part of the stratum moleculare of the proximal Sub, whereas the ventral part of A35 projects to the deep part of the stratum moleculare of the distal Sub. Furthermore, it has also been reported that rostral and caudal parts of A35 both project to septal and temporal levels of the Sub. Supplement 2 shows that this information is based on two references 1-2. References:

1. T. W. Deacon, et al., "Afferent connections of the perirhinal cortex in the rat," J. Comp Neurol. 220(2), 168 (1983).
2. S. C. Furtak, et al., "Functional neuroanatomy of the parahippocampal region in the rat: The perirhinal and postrhinal cortices," Hippocampus (2007).
3. K. C. Kosel, G. W. Van Hoesen, and D. L. Rosene, "A direct projection from the perirhinal cortex (area 35) to the subiculum in the rat," Brain Res. 269(2), 347 (1983).

Quality assurance

References of all connections are provided in Supplement 2. However, for each connection that is drawn, we can provide the source reference(s) and the page and/or figure number in which it was described. Most questions regarding the literature can be answered by looking into the reference lookup table. In case you require more details about the references than provided in Supplement 2, we kindly invite you to send your questions by email. Unfortunately we cannot distribute original articles, since they are copyright protected. Should you feel that a connection is missing, please send the details of the reference in which the connection is mentioned. We will review the suggestion and if accurate, we will add it to our diagram. Although much care has been taken to make this diagram as accurate as possible, small inaccuracies may exist in the almost 1600 connections that were hand-drawn for this diagram. Should you find errors or have suggestions to improve the diagram, please send an e-mail.

Thank you!

Appendix 1: Abbreviations

• A35: Area 35 of the perirhinal cortex
• A36: Area 36 of the perirhinal cortex
• CA1: Cornu Ammonis 1
• CA3: Cornu Ammonis 3
• caud: caudal
• crest: crest area of DG
• DG: Dentate Gyrus
• dist: distal part
• dl: dorsolateral band of the entorhinal cortex
• encl: enclosed blade of DG
• exp: exposed blade of DG
• gran: granular cell layer
• hilus: polymorphic layer of DG
• I, .., VI: first to sixth layer of the isocortex of the parahippocampal area
• im: intermediate band of the entorhinal cortex
• iml: inner 1/3th of the molecular layer
• int: intermediate part, i.e. in between the proximal and distal part
• LEA: Lateral entorhinal area
• luc: stratum lucidum
• MEA: Medial entorhinal area
• ml deep: deep part of the molecular layer
• ml sup: superficial part of the molecular layer
• mml middle: 1/3th of the molecular layer
• oml outer: 1/3th of the molecular layer
• or: stratum oriens
• PaS: Parasubiculum
• POR: Postrhinal cortex
• prox: proximal part
• PrS: Presubiculum
• pyr: pyramidal cell layer
• rad: stratum radiatum
• rost: rostral
• slm: stratum lacunosum moleculare
• Sub: Subiculum
• unsp: unspecified
• vm: ventromedial band of the entorhinal cortex

Appendix 2: Problem solving

Problem: Sometimes Adobe 8 shows a "content preparation" dialog box, which indicates that the software is preparing the document in some way for reading. Unfortunately, this dialog returns every time you switch on/off a layer in the layer navigation pane. This slows down the interaction with the diagram to an undesirable speed.

Cause: The problem is caused by a screen reader plugin that is automatically loaded on Adobe 8 startup.

Solution: This plugin cannot be disabled from the Adobe 8 interface. Therefore one has to manually disable the plugin. To do so, follow these steps:

  1. Close Adobe Reader
  2. Open Windows explorer and navigate to the plugins subdirectory of the Adobe Reader install directory (by default C:\Program Files\Adobe\Reader 8.0\Reader\plug_ins\)
  3. Find the following files in these folder: Accessibility.api, MakeAccessible.api, ReadOutLoud.api
  4. For safety, store them in a zip archive. This will allow you to undo the action.
  5. Delete these 3 files.
  6. Restart Adobe
  7. Select "OK" on any message dialog that complains about loss of functionality.

Applicability: This fix has been tested only on Windows XP (SP3) with Adobe Acrobat Reader version 8. However, it is likely that this solution will also work for Acobat Reader 9. Please feel free to provide us with feedback, so that other users may benefit from your experience.