Flashcards
Learning - O+E
Flashcards do conjunto (23)
- 1Learning = acquisitor of altered behavioural response due to an environmental stimulus Memory = process through which learned info is stored and retained Recall = conscious or unconscious retrieval process by which altered behaviour is shownMemory:
declarative = facts & events
non declarative = skeletal musculature, emotional response - 2procedural learning = learning a motor response in reaction to a sensory input procedural memories: - can be formed along simple reflex pathways - amendale to investigation 2 types: non-associative & associative learningnon-associative
- change in behavioural response that occurs over time in response to a single type of stimulus
2 types: habituation & sensitisation
habituation = learning to ignore a stimulus that has meaning
sensitisation = strong stimulus results in exaggerated response to all subsequent stimuli - 3Associative learning - 2 types: - classical & instrumental conditioning classical: associating a stimulus (unconditioned) that evokes a measurable response with a second stimulus (conditioned) that doesn't normally evoke this response - learned response = conditioned response - timing of US & CS is very importantInstrumental conditioning:
- learning to associate a response (motor act) with a meaningful stimulus
- timing is important
- motivation plays key role (underlying neuronal circuits are more complex) - 4Non-associative learning in Aplysia - habituation & sensibilisation of gill with drawl reflex gill withdrawal reflex: - if mantle removed, underneath is gill - spray siphon with water, sensory stimulus causes siphon & gill to withdraw - at motor neurone, ESPS will decline over time -> proves change occurs at synapsesensitisation of gill withdrawal reflex:
- if stimulate head at same time as siphon, response is enhanced
- PKA phosphorylates potassium channels -> K channels close -> prolonged AP -> more Ca entry -> more NT (quanta) released
- release of NT serotonin binds to couple receptors activating cAMP which vitiates PKA - 5glutamte contributes to persistent sensitisation - delivery of new glutamate receptors to post-synaptic membraneassociative leaning in Aplysia -> classical conditioning:
- involves 3 neurona circuits + paired stimuli
- spray of water = gentle, not huge response
- increase calcium helps produce more cAMP therefore more response in post-synaptic cell - 6invertebrate studies: - modifications in synaptic transmission results in learning & memory - changes in neuronal activity can modify intracellular second messenger pathways - alterations in existing synaptic proteins can result in learning & memorySynaptic plasticity:
- widespread in CNS of vertebrates
- different types of synapses have slightly different mechanisms underlying plasticity
- mechanisms of plasticity best understood in cerebellar cortex & hippocampus - 7Cerebellum (back of brain) - important site for motor learning - can correct movement (not initiate)cerebellar cortex
major inputs to purkinje cells - receive axons from 2 cell types:
- parallel fibres (arising form cerebellar granule cells)
- claiming fibres (arising form inferior olive)
major input to granule cells:
- mossy fibres (airing from neurones in pontine nuclei)
purkinje cells activated - release inhibitory neurotransmitter - 8cerebellar long-term depression (LTD) - record electrical signals from cells & infer whats changing & underlying signals are - put electrode into purkinje & record response in response to stimulus - if stimulate climbing fibres same time as purkinje, cell receives two inputs (causes change in response) long term depression -> decreasing response = basis for learning in cerebellummechanism of LTD induction in cerebellum
- glutamate released from parallel fibre terminals bind to mGluR1 at perisynpatic region of purkinje dendrite spines - 9Hippocampus - declarative memory involved neocortex & structures in medial temporal lobe in humans, hippocampus is: - important for spatial memory & word recall - implicated in several clinical disordersright hippocampus = active during learning & recall of spatial environments
left hippocampus = domain for word recall
- region of temporal lobe removed =lose ability to form new memories
- proves hippocampus important in forming memory - 10Hebb's postulate - axon of cell A near enough to excite cell B & repeatedly takes part in firing it, growth process/metabolic change takes place in one/both cells such that A's efficiency is increased - cells that fire together, wire togetherLong-term potentiation (LTP)
- stimulation of axons at low basal rate produce stable synaptic response
- application of single high frequency stimulus train resulted in persistent increases in response size - 11Hippocampus plays role in spatial learning = trisynaptic circuit - very layered regions of cells CA3 cells release neurotransmitter to CA1 cellselectrophysiological recordings:
- extracellular
- intracellular (single channel or whole cell recording)
- in vivo recording - 12NMDA receptors - multimeric complex of different subunits - ionotropic receptor binds glutamte - glycine is co-agonist - Ca2+/Na+ channel - magnesium blocks pore unless depolarisedexpression of LTP
- increase in postsynaptic responsiveness
- post translational modification of AMPA receptors
- increased number of AMPA receptors
- activity-dependent exocytosis & lateral diffusion from extra synaptic sites (2 mechanisms for delivery)
- long term change due to changes in post-synaptic cell (enables more ample receptors in membrane, more ion channels to respond to glutamate) - 13Hippocampus - 3 cell types: - CA1 pyramidal cell - CA3 pyramidal cell - dentate gyrus granule cellelectrophysiology
- put stimulating & recording electrodes in appropriate places to stimulate neurones to induce LTP or LTD - 14long-term potentiation (LTP) - recording electrodes = glass pipettes & fill with solution (mimics ECF) - have control to get base line responses - 2 stimulating electrodes, each on diff axons, record from one cell - apply slow stimulation to one electrode, response in recording should stay at base line - in other pathway: have slow stimulation then high frequency stimulation (if enough to affect synapse, will get LTP)induction of LTP: role of NMDA
- LTP induced by high frequency stimulation
- sudden high frequency stimulation burst of activity enough to release glutamate (causes change in post-synaptic cell - makes more responsive to simulation)
- burst of stimulation opens NMDA receptors to allow for Ca entry - 15Preparing for dissection - make sure everything sterile - vibratome = experimental machine makes cerebral spinal fluid 1. remove brain & put immediately in artificial CSF & oxygenate 2. vibratome has shape razorblade & moves across tissue making v fine sections (400um thick) 3. collect brian slices, suck up with pipette & transfer to dish *mount in order to cut right section instead of killing cellspipettes prepared by putting in heated ring to form thin glass pipette
- need to know ohms law for electrophysiology V=IR
- pipette hole must be appropriate size to attach to cell (too big will suck up cell) - 16Need chamber: - place slice in solution used to being in (osmolarity must be accurate) - can control speed of flow - keeps brain slice alive - O2 take feeds in oxygen aCFS = artificial cerebral spinal fluid (95% O2, 5% CO2)making solution:
- if incorrect sample will die
- concentrations, osmolarity & pH
- used double distilled water
intracellular recordings:
- record form one pyramidal cell
- at threshold, fire APs - 17two types of brain slices: acute = slices prepared fresh each day organotypic = slices prepared from younger animals - slices are cultured*working environment must be sterile
- wear gloves & face mask
- pray ethanol on everything
- use cell culture hood (all contaminants sucked away)
- use para-film -> seals containers so nothing can come in bottles & contaminate
- incubator to stop organotypic brain slices (keep at correct temp)
- cell future media must be changed frequently
- exposes slices as little as possible - 18Laser scanning microscopy - laser light source - laser beam needs to be rapidly scanned to illuminate entire visual field - photomultiplier (PMT) measures fluorescence detected at each pointconfocal laser scanning microscopy
- pinhole removes out-of-focus background fluorescence
- good for imaging relatively thin samples - 192-photon laser scanning microscopy - one pulsed laser - deeper imaging (IR light) - low level of photo-toxicity - minimal photo-bleaching - reduced light scatteringbiolistic transfection of organotypic slices
- gets fluorescent probe into brain slice
- use gene gun & shoot slices with bullets
- can make hole in cell & inject fluoresce right into cell (alternate technique) - 20dynamic calcium investigation: - looking at agonist & antagonist (active/inactive receptors) - different techniques for different neuronal preparationsusing fluorescence to detect changes in calcium - calcium:
- important second messenger in neuronal signalling
- controls activity of many enzymes
- plays important roles in membrane fusion
- important for synaptic plasticity
(more calcium, brighter the dye will be) - 21getting calcium indicator into cells: - load individual cells using whole-cell patch-clamp technique - inject calcium indicator into individual cells using sharp electrodesuse AM-version of dye -> esterified molecule (can diffuse across membrane) -> once inside cell is de-esterified inside neurone by enzymes
- cells then washed to removed remaining dye outside cell (when excited with light, cell will fluoresce) - 22Acute slice - advantages: - can make intra/extracellular recordings from cells - neuronal activity closely reflect in viv situation - fresh tissue so little chance of infection - easy to obtain clean recordingsdisadvantages:
- thick slices
- difficult to get good images with confocal
- 2-photon microscopes v expensive
- rarely use all slices you prepare
- reduced experimental time - 23Organotypic slice - advantages: - slices kept alive in incubator for weeks - use all tissue you prepare - can combine with transfection techniques - slices are thin -> compatible with confocal imaging - less material wasted as several experimenters can use slices from one animalDisadvantages:
- need to prepare in sterile environment (prone to infection)
- must change media every 2 days
- not as representative of in vivo (new pathways can form)
*both preparations:
- can take slices from transgenic animals
- can make complete brain slices
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- FlashCards sobre Learning - O+E, criado por michaela minkin em 20-05-2018.
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- michaela minkin
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