1
Q
How to Measure Brain Connectivity:
How do you see neurons?
A
In order to see neurons you need contrast
- Golgi Stain Method
- fills cells with silver-chromate
- Fluorescent dyes (GFPs)
- GFP for brain labelling
2
Q
What are the drawbacks to Golgi stain method and Fluorescent dyes?
A
Both techniques require neurons to be traced out manually or with software
3
Q
What is the difference between Anterograde Tracing neuron connections and Retrograde tracing?
A
- Anterograde tracing neural connections
- Carry dyes/fluorescent proteins through axons to be visualized
- Dyes, viruses (adenoassociated viruses (AAVs))
- Retrograde tracing
- Dye or compound gets incorporated into the axon and travels backward to the cell body
- Cholera toxin, fast blue. Adenoassociated viruses (AAVs), Rabies
4
Q
What are four techniques to stimulate neurons?
A
- Electrical stimulation
- Light stimulation (optogenetics)
- Chemical stimulation (pharmacogenetics)
- Patch clamp single cell stimulation
5
Q
Electrical Stimulation:
- Technique
- Pros/Cons
A
Electrical Stimulation:
- Technique
- place wire into brain tissue
- pass current into tissue to depolarize neurons near the electrode
- Record from another brain region to see if neurons respond to stimulation
- Pros/Cons
- Pros
- easy to implement
- effective (repeatable)
- Precise activation onset
- Cons
- Indirect, unintended activation of other neurons close to the stimulation electrode
- antidromic activation of post-synaptic cells
- ie no specificity to which region is responding to stimulation
- Pros
6
Q
Optogenetic Stimulation
- Technique
- Pros
- Cons
A
Optogenetic Stimulation
- Technique
- light sensitive rhodopsin is genetically expressed in neurons of interest
- In the presence of light, cells are depolarized and can be activated
- Pros
- Rapid control of spike timing
- specific (genetically defined) neuron types can be activated without unintended activation of nearby neurons in the brain
- Cons
- Light can change the temperature of neural tissue
- Must deliver light to the brain using brain implants
7
Q
What is the first light dependent depolarization opsin used to activate neurons?
A
Channelrhodopsin (ChR2) - responds to blue light
8
Q
Chemogenetic stimulation
- Technique
- Pros
- Cons
A
Chemogenetic stimulation
- Technique
- Designer (bioengineered) receptor is expressed in cells of interest using genetic approaches
- Receptor is designed to be activated by a specific ligand (drug)
- Pros
- cells can be activated by simply applying a drug
- Drug acts specifically on the designer receptors
- Specific cell types can be activated
- Cons
- No precise control over the timing of activation
- ie drug action can be long
- No precise control over the timing of activation
9
Q
Paired Patch Clamp Recordings
- Technique
- Pros
- Cons
A
Paired Patch Clamp Recordings
- Technique
- 2 single neurons are recorded using intracellular techniques (so that they can be depolarized with current injection)
- Pros
- Definitive test of connectivity between neurons in the brain
- Only true physiological way to test connectivity between neurons
- Cons
- Challenging to implement (hard technique)
- High failure rate
- Only useful for testing close connections
10
Q
Cortical Connectivity: micro vs macro
- Basic Connectivity Rules
- Within brain region
- Between brain regions
- Long range
- Micro connections
A
Cortical Connectivity: micro vs macro
- Basic Connectivity Rules
- Within brain region
- Cells close to each other are more likely to connect to each other
- Between brain regions
- there is no (or weak) relation between distance and connectivity
- Long-range connections
- Majority are excitatory
- Micro connections
- majority are inhibitory
- Within brain region
11
Q
What are the two neuron types?
A
- Excitatory - pyramidal cells
- Inhibitory cells - Interneurons
12
Q
- Excitatory - pyramidal cells
- release ______
- action
- Morphology
- Projection
- Inhibitory cells - Interneurons
- release _____
- action
- morphology
- projection
A
- Excitatory - pyramidal cells
- release Glutamate
- action:
- excite post synaptic cell
- Morphology
- larger in diameter
- many dendritic spines
- Projection
- project locally to nearby cells and to different regions of the brain
- Inhibitory cells - Interneurons
- release GABA
- action
- inhibit post-synaptic cell
- morphology
- smaller in diameter
- generally lacking spines
- projection
- mainly project locally within 0.2mm (recieve input from other regions but inhibit locally)
13
Q
What are the four interneurons and their connections?
A
- Parvalbumin (PV)
- Synapse on cell bodies
- GABA-A mediated inhibition
- Somatostatin (SST)
- Synapse on dendrites
- GABA-A mediated inhibition
- Vasoactive intestinal polypeptide (VIP)
- synapse on other interneurons
- GABA-A mediated inhibition
- Neuropeptide Y/Neurogliaform cells (NG)
- express nitric oxide synthase and Neuropeptide Y
- Synapse on other interneurons AND excitatory cells
- GABA-B and Volume transmission
14
Q
Label the types of inhibition represented by colour
A
Blue: Feedforward inhibition
Red/Blue: Feedback inhibition
Red: Lateral inhibition
Green: Disinhibition
Yellow: Volume inhibition
Black: Feedforward excitation
15
Q
What is feedforward inhibition?
What type of neurons typically mediate feedforward inhibition
Purpose?
Gone wrong?
A
- Inputs activate interneurons (without necessarily activating pyramidal cells)
- PV (parvalbumin) cells mediate feedforward inhibition
- Feedforward inhibition acts to filter inputs
- With no PV cells, pyramidal cells fire in excess, similar to epilepsy
16
Q
Feedback inhibition:
- Definition
- Type of interneuron
- Purpose
A
Feedback inhibition:
- Definition
- when excitation in one cell activates an interneuron to generate inhibition of itself
- Type of interneuron
- PV and SST cells can both participate in feedback inhibition
- PV=Parvalbumin
- SST = Somatostatin
- PV and SST cells can both participate in feedback inhibition
- Purpose
- Provides stability (prevents excess excitation)
- FB inhibition arises from local excitation in neural circuits (in contrast with FF inhibition which can be generated by long range excitation)
17
Q
Lateral Inhibition:
- Definition
- Interneurons
- Purpose
A
Lateral Inhibition:
- Definition
- one pyramidal cell activates an interneuron to inhibit another pyramidal cell
- Interneurons
- SST cells mediate lateral inhibition
- W/o SST cells, other pyramidal cells can fire when they’re not supposed to
- Purpose
- Generates segregation btwn neural groups
- Allows firing of one cell to stand out
18
Q
Disinhibition
- Definition
- Interneurons
A
Disinhibition
- Definition
- inhibit inhibitory cell connectivity creating net excitation of another cell population
- Interneurons
- VIP→SST connection is disinhibitory with respect to pyramidal cells
- Without VIP (vasoactive intestinal polypeptide) cells, pyramidal cells fire less
19
Q
Volume inhibition
- Definition
- act at
*
A
Volume inhibition
- Definition
- Neurogliaform cells release GABA and act on GABA-B receptors and GABA-A (slow) receptors
- Effects can be synaptic & extrasynaptic
- NG cell can therefore influence activity in a non-synaptic way
20
Q
Feedforward excitation
- Definition
- Connections
A
- Definition:
- Excitatory cells mediate excitation of other neurons
- An essential feature of communication within and between brain regions
- Connections between pyramidal cells is 3-10% so connectivity is relatively infrequent
- however, most neurons are excitatory so a small number of pyramidal cells can exert a lot of excitation
- Feedforward excitation is usually specific: neurons that “code” for 1 stimulus will communicate
21
Q
Feedforward excitation
- Definition
- Connections
A
- Definition:
- Excitatory cells mediate excitation of other neurons
- An essential feature of communication within and between brain regions
- Connections between pyramidal cells is 3-10% so connectivity is relatively infrequent
- however, most neurons are excitatory so a small number of pyramidal cells can exert a lot of excitation
- Feedforward excitation is usually specific: neurons that “code” for 1 stimulus will communicate
22
Q
What is meant by balanced excitation and inhibition?
A
- The thalamus excites layer 4 of the visual cortex
- Interneurons are also excited by thalamic input
- Inhibition strength and excitation strength are correlated
- Small increases in the excitation:inhibition ration generates APs
23
Q
When you move, you generate “self-generated sounds”. How does the brain tune-out these sounds in order to allow other sounds to be heard while moving
A
- Feedforward inhibition
- Motor cortex is activated during movement (eg running)
- Excitatory neurons from motor cortex activate PV interneurons in auditory cortex
- PV interneurons suppress auditory cortex during movement
- Free up space for external stimuli
24
Q
- Gamma rhythms can be recorded from the _____ during periods of _______
- They require the activity of ______
- _______→_______ sequences, usually give rise to _____ activity
- Decay of inhibitory potentials can control the _______ of firing in ______
A
- Gamma rhythms can be recorded from the cortex during periods of increased attention
- They require the activity of both excitatory (pyramidal) cells and interneurons
- excitation→inhibition sequences, usually give rise to rhythmic activity
- Decay of inhibitory potentials can control the frequency of firing in pyramidal cells
25
* What type of inhibition participates in Gamma rhythm generation?
* What stimuli can evoke gamma rhythms in the visual cortex
* Suppressing ______ decreases gamma rhythms
*
* What type of inhibition participates in Gamma rhythm generation?
* **feedback and feedforward inhibition**
* What stimuli can evoke gamma rhythms in the visual cortex
* **visual stimuli**
* Suppressing _SST cells_ decreases gamma rhythms
* suggests feedback inhibition plays a role
* Feedforward inhibition also plays a role in generating gamma rhythms in other neural structures like the hippocampus
26
Surround suppression (lateral inhibition) in the visual cortex:
* Pyramidal cells in V1 ______ their firing with larger stimuli than outside their receptive field (surround suppression)
* _____ interneuron increases their activity
* Surround suppression arises from \_\_\_\_\_\_\_
* May participate in ability to \_\_\_\_\_\_\_
* Pyramidal cells in V1 decrease their firing with larger stimuli than outside their receptive field (surround suppression)
* SST interneuron increases their activity
* Surround suppression arises from **lateral inhibition**
* May participate in perceptual ability to recognize continuity of objects in the visual field
27
Disinhibition (brain state - locomotion)
* During locomotion, ____ become active even with no visual stimuli
* ____ cells are activated by acetylcholine which induces \_\_\_\_\_
Disinhibition (brain state - locomotion)
* During locomotion, _V1 cells_ become active even with no visual stimuli
* _VIP_ cells are activated by acetylcholine which induces _disinhibition of pyramidal cells_
VIP = vasoactive intestinal polypeptide (VIP) interneurons
28
What is the role if VIP and SST in control of blood flow?
* VIP induces vasodilation, increasing blood flow (increase excitation to increase blood flow to active regions)
* SST induces vasoconstriction, decreasing blood flow
* Similar opposing actions at the level of blood flow and regulating excitability in pyramidal cells
VIP increases → SST decreases → excitation increases
29
What is global inhibition? What gene is associated and which interneurons show that gene
Sleep
* c-fos is an immediate-early gene that indicates neurons with high AP firing rates
* Following sleep, NG cells show c-fox in the cortex
* Other cell types do not show c-fox
* NG cells may cause decreased cortical activity during sleep
30
What four processes all require unique contributions from different types of neurons as discussed in class?
Gamma rhythms
Locomotion
Auditory processing
Visual coding
PHYSL 372 - Systems Neuroscience
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