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C. elegans neuronal network (279 out of 302 total neurons)
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Determined through tracing and electron microscopy
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red: sensory neurons, blue: interneurons, green: motor neurons
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6393 chemical synapses, 890 electrical junctions, and 1410 neuromuscular junctions
Anatomical connectivity
White et al, Phil Trans R Soc Lond B 1986
http://www.wormatlas.org/neuronalwiring.html
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Wiring diagram of C. elegans (302 neurons total, 279 neurons shown here. 20 pharyngeal (of or relating to the pharynx) nervous system neurons not shown as well as 3 that do not make synapses with other neurons)
red: sensory neurons
blue: interneurons
green: motor neurons
number of possible node pairs is N*(N-1)/2
for c. elegans: > (302*301)/2
[1] 45451
(279*278)/2
[1] 38781
6393 chemical synapses, 890 electrical junctions, and 1410 neuromuscular junctions
horizontal axis represents closeness of connectivity (spring embedded graph layout)
vertical aix represents signal flow from top to bottom
from D. Chklovskii. White et al, Phil Trans R Soc Lond B 1986
Genome size does not correlate with nervous system complexity
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100,000,000,000
71,000,000
302
10,000,000
250,000
Number of
neurons in whole nervous system
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Number of genes is not related to nervous system complexity or size. The nematode c. elegans has just 302 neurons, and yet its genome contains virtually as many genes as a humans. An african elephant brain weighs 3 times more than a human brain and has 3 times the number of neurons.
The largest brains are those of sperm whales, weighing about 8 kg (18 lb). An elephant's brain weighs just over 5 kg (11 lb), a bottlenose dolphin's 1.5 to 1.7 kg (3.3 to 3.7 lb), whereas a human brain is around 1.3 to 1.5 kg (2.9 to 3.3 lb). Brain size tends to vary according to body size.
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Investigating gene function
Principles of Neurobiology, Garland Science Fig. 13-4
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Forward and reverse genetics.
forward genetics, researchers start by observing an altered trait (phenotype) to id. the gene responsible for causing the phenotype of interest.
in reverse genetics, researchers start with a gene of interest and disrupt the gene function, examing the phenotypic consequences.
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Making mammalian genetic models
Principles of Neurobiology, Garland Science Fig. 13-6
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Making fancy mice.
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- CRISPR/Cas9 system for faster genetic engineering
Targeted editing of the genome
Principles of Neurobiology, Garland Science Fig. 13-8
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crispr-cas9 system.
crispr: clustered reguarly interspaced short palindromic repeat
cas: crispr associated
any eukaryotic dna that contains a PAM sequence can be a target.
PAM sequence: protospacer associated motif, usually two or three nucleotides. Occurs frequently.
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-canonical PAM is the sequence 5'-NGG-3' where "N" is any nucleobase followed by two guanine ("G") nucleobases
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-Guide RNAs (gRNAs) can transport Cas9 to anywhere in the genome for gene editing, but no editing can occur at any site other than one at which Cas9 recognizes PAM.
A guide RNA that contains sequences complementary to a piece of DNA from the target gene of interest brings the Cas9 enzyme to the target site on the chromosome through DNA-RNA base pairing (purple and red)
Two nuclease domains of Cas9 create a double strand break in the genomic DNA. The double strand break can be repaired by the nonhomologous end joining system, through which small deletions or insertions may be created at the repair site.
The ds break can also be repaird by the homologous recombination system with a donor DNA as a template, through which specific modifications such as insertion of a transgene can result.
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Timeline of genome sequencing advances
Principles of Neurobiology, Garland Science Fig. 13-7
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timeline of selected milestones. Graph show exponential growth of sequencing technology in 10 yrs since draft of human sequences first published.
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Traditional anatomical methods for determining cortical areas
Principles of Neurobiology, Garland Science Fig. 13-18
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Nissl stain and cortical area divisions. Border of V1 and V2. Adapted from Brodmann K (1909).
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Confocal and light sheet fluorescence microscopy
Principles of Neurobiology, Garland Science Fig. 13-19
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confocal is on left. light sheet microscopy on the right.
Confocal microscopy uses a small pinhole before the detector to allow emitted fluorescence from only the focal plane.
In light sheet microscopy, illumination is proved from the side to produce a thin sheet of excitation light.
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Making brain tissue transparent for better microscopic imaging
Principles of Neurobiology, Garland Science Fig. 13-20
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clarity based tissue clearing for fluorescene imaging.
Intact tissue is fixed in prescence of hydrogel monomers that covalently link DNA RNA, and proteins into a mesh during subsequent polymerization. Lipids (which are a major cause of opacity for fluorescence imaging) are not covalently linked and are removed during subsequent clearing process by passive diffusion or electrophoresis in presence of detergent. Tissue is then transparent for better and deeper imaging. This example is from a Thy1-gfp transgenic mouse imaged with a confocal microscope. Shows neocortex, hippocampus, and thalamus.
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Injecting intracellular dyes to trace axonal projections
Principles of Neurobiology, Garland Science Fig. 13-21
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Intracellular dye filling to trace axonal projections of a single neuron. Rat posterior piriform cortex pyramidal neuron. Injected in vivo followed by several days. Johnson et al., J Neurosci 2000.
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Super-resolution fluorescence microscopy
Principles of Neurobiology, Garland Science Fig. 13-25
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Mapping synaptic protein organization with super res fluorescence microscopy.
double labeling with basson (presynaptic scaffold protein, red) and postsynaptic scaffold protein (homer1, green) in mouse olfactory bulb glomerular layer.
Imaged using stochastic optical reconstruction microscopy (STORM), a super resolution technique. Gets you beyond the diffraction limit for light microscopy, typically 100-150 nm with the highest resolution objectives and shortest wavelengths of visible light.
Third image is higher magnification view.
Right shows distribution of different antibody, STORM imaging resolved proteins around the synaptic cleft.
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Diffusion tensor imaging
Principles of Neurobiology, Garland Science Fig. 13-26
www.humanconnectomeproject.org
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Diffusion tensor imaging.
sagittal view of human brain.
Axon bundles running along medial-lateral axis are colored red.
Those along anterior-posterior axis colored green.
Axons running through brainstem colored blue.
www.humanconnectomeproject.org
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Tracing of long distance connections in the mouse brain
Principles of Neurobiology, Garland Science Fig. 13-27
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Mix of phytohemagglutnin (PHA-L, green) an anterograde trace and cholera toxin subunit b (CTb, magenta) a retrograde tracer injected into right insular cortex of a mouse brain. Stained in blue with fluorescent nissl stain. Projections from and to insular cortex.
On right show adeno-assoc viruses expressing Cre-dependent GFP. Injected into motor cortex of mic expressing Cre recombinase in layer 6 or 2/3. Notice different projection patterns of neurons from these two layers in the rendered 3D views.
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Reconstruction of wiring diagram using serial EM
Principles of Neurobiology, Garland Science Fig. 13-29
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serial electron microscopy to construct wiring diagrams.
left is EM micrograph of drosophila medulla in optic lobe.
High mag view shown at bottom showing presynaptic terminal with 4 contacts.
Colored view showing segmented cellular elements in this image.
Neurites reconstructed by registering and linking thousands of consecutive brain sections… from Takemura et al, Nature 2013
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Using modified rabies virus to do monosynaptic tracing
Principles of Neurobiology, Garland Science Fig. 13-30
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Gene encoding rabies virus glycoprotein is essential for viral recognition of host cells and for viral spread is replace with GFP in the rabies genome. This mutant rabies virus can no longer recognize and transduce normal mammalian neurons.
Mutan rabies assembled in a cell line that helps assemble the virus with a coat protein from a different virus, the EnvA coat protein (blue). This makes it able to transfect mammalian cells that express the EnvA receptor TVA (cyan) from a transgene.
A transgene that supplies the rabies glycoprotein is alos expressed in the starter cells.
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Principal methods for electrophysiology
Principles of Neurobiology, Garland Science Fig. 13-31
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left is photo uncaging
right is optogenetics
parse out signal flow in the brain.
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Multielectrode arrays
Principles of Neurobiology, Garland Science Fig. 13-33
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multielectrode arrays (like this 10x10 silicon based prototype) are widely used now for recordings from multiple cortical neurons simultaneously and for usages with neural prosthetics. from IEEE 1991
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Laser scanning two-photon microscopy: imaging neurons in living mice
Principles of Neurobiology, Garland Science Fig. 13-39
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two photon microscopy is non-linear magic.
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Chemical and genetically encoded calcium indicators
Principles of Neurobiology, Garland Science Fig. 13-38
GCaMP6– genetic calcium indicator
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chemical and genetically encoded calcium indicators.
fura2 is a fluorophore with a calcium chelating site from calcium buffer EGTA. With low calcium, excitation at 380nm produces stronger fluorescence emission than excitation at 350 nm. Ratiometric imaging at 350/380 gives sensitive measure of [Ca2+]
first genetic calcium reporter based on FRET (fluorescene resonance energy transfer).
Gcamp. permutated gfp is restored to its native 3D structure with an associated increase in fluorescene after calcium triggered binding of the calmodulin binding peptide M13 to calmodulin. Fluorescent intensity thus gives readout of Ca2+. Single APs reliably induce fluorescent intensity changes in GCaMP6 in mouse visual cortical neurons in vivo.
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Recording neuronal activity in awake behaving mice
Principles of Neurobiology, Garland Science Fig. 13-40
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Head fixed preparation on left during 2P imaging. The thirsty mouse can be trained to extend its tongue only when odor A, but not odor B is presented in order to receive a water reward. Motor cortical area controling tongue extension can be imaged during the learning process.
Virtual reality preparation to test the neuronal correlates of memory dependent spatial navigation. Floating styrofoam ball that the mouse is trained to run on, head fixed with electrodes or 2P imaging. Screens providing continuous first person VR experience— in fact this was done by coding a VR environment based on the quake 2 game engine (if any of you have played the classic quake first person shooter games by id software).
The right shows a miniaturized microscope weighing just over a gram that an adult mouse can carry on its head to image brain activity as it freely moves and navigates.
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Optical imaging of brain activity
🎥
💭
- Record brain activity patterns
J. Ackman
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use modern CMOS cameras and transgenic mice to image brain activity across both cerebral hemispheres simultaneously so that we are covering fields of view of 10-20mm.
And this is now possible because we can use new transgenic mice that express a green fluorescent reporter protein that specifically exhibits increased fluorescence when neurons are electrically active. And what we do is to record population activity patterns transcranially with what we call functional mesoscale optical imaging.
Mesoscale imaging of neocortical dynamics
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1 mm
J. Ackman
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And so here is the field of view of our preparation and what you’re looking at is the two cerebral cortical hemispheres of a young mouse being that was imaged transcranially.
Mesoscale imaging of neocortical dynamics
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11x13mm FOV, postnatal day 5 (P5), SNAP25-GCaMP6
1000 µm
J. Ackman
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And if we want to know what population activity looks like across the developing cortex, here is your answer…
that population activity in the developing cortex consists of discrete but overlapping domains of activation. So what you are seeing is the ongoing neuronal population activity throughout localized parts of the neocortex in a head stabilized, unanesthetised and resting young mouse. These movies are 10 min long recordings, that are acquired at a framerate of 5-10 fps, played back 6x here, and has been normalized to the mean fluorescence image for the entire 10 min long movie so that you are looking at changes in pixel intensity. This means you see neuronal activity show up as these bright blobs in the brain, as well as motion of the paws, hair, and whiskers of the animal within the field of view.
We think much of this activity is spontaneous but some of it is self stimulated tactile responses because the animals often exhibit limb twitches just as you do when you’re drowsy or when you were in the womb.
This technique is important because the functional organization —> of the mature brain is distributed across areas with different functions within and between hemispheres.
Because this method is minimally invasive, requiring just retraction of the scalp and no local dye injections, this will be a technique of choice for large range of experiments including those requiring in vivo imaging in different behaving, unanesthetised mouse lines.
And to illustrate the sensitivity of this cortical activity to general anesthesia, we can watch the following movie…
Optogenetics – a new technique for understanding brain function
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http://www.youtube.com/watch?v=I64X7vHSHOE
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Channelrhodpsin-2 (ChR2) from green algae is a cation channel gated by blue light
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Allows more Na+ influx than K+ efflux causing depolarization
Optogenetics for precise control of neuronal activity
Principles of Neurobiology, Garland Science Fig. 13-45
http://www.youtube.com/watch?v=I64X7vHSHOE
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- Combining methods for read/write control— of the brain!
Mapping connectivity by combining optical and electrophysiological methods
Principles of Neurobiology, Garland Science Fig. 13-47
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left is photo uncaging
right is optogenetics
parse out signal flow in the brain.
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- Multimodal sensory activation, focuses attention, object tracking and stereoscopic vision/depth perception, salience of object, defensive/self preservation, limbic loop, decision making, movement selection, proximal and distal motor pool recruitment, reflex activation— but all from a previously unexperienced activity!
“I didn’t have time to think”
2016-03-10 09:48:33
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- 30 ms before reaching LGN. 60 ms to primary visual cortex. Color information slower (M cell vs P cell pathways).
Response latency in macaque visual system
2016-03-10 09:48:33
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cell body (soma)– metabolic center of the cell, contains the nucleus.
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dendrites– receive incoming signals from other nerve cells
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axon– carries signals to other neurons
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axon hillock– initiates action potentials
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synapse– site at which two neurons communicate
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synaptic cleft– area between pre and post-synaptic cell
Structures of a neuron
2016-03-09 11:31:59
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Neuron Processes: Action Potentials
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Nerve impulse (action potential)
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Neuron receives and sends signals
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Generated at the initial segment of the axon
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Conducted along the axon
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Releases neurotransmitters at axon terminals
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Neurotransmitters – excite or inhibit neurons
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Neurons communicate by electricity
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Axons project great distances
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Neurons do not touch each other directly.
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Come in close proximity at the synapse
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Use action potentials to transmit information
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Action potential causes release of neurotransmitter that is received by post-synaptic cells.
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Ways to measure neural activity
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Extracellular recording– an electrode is placed near a neuron. Measures action potentials. Useful for detecting patterns of activity.
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Intracellular recording– an electrode is placed inside a neuron-can measure smaller graded potential changes. Useful for isolating responses to single inputs.
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Voltage clamping– can make the membrane potential of a cell at a desired point and determine the current flow across the membrane.
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Patch Clamping– can measure ion flow across a single channel.
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fMRI– infer activity indirectly in a living brain based on brain oxygenation patterns
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Basic parts of the CNS
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Spinal cord
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Brain stem
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medulla
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pons
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midbrain
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Cerebellum
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Forebrain
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diencephalon
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cerebral hemispheres
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These are the basic parts of the CNS
Words used to describe locations in the CNS
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General rules of spinal cord organization
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Neurons and axons that process and relay sensory information (afferents) are in dorsal spinal cord.
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Preganglionic visceral motor neurons (innervate glands) are found in the intermediate/lateral region.
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Interneurons are in intermediate zone.
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Motor neurons and axons are found in the ventral portion of the cord.
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Distal muscles are innervated by lateral motor neurons.
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Proximal muscles by medial motor neurons.
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Spinal cord tracts
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Dorsal column– sensory signals travels up it to the brain.
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Lateral columns– also called the cortico-spinal tracts. Take signals from brain and sends it to the muscles.
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Ventral columns (sometimes called anterolateral column)– carry pain signals up and motor signals down.
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Spinal cord tracts
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Nissl stain (cell bodies)
Myelin stain
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Dorsal column-sensory info travels up to the brain.
Lateral columns-also called the cortico-spinal tracts. Take info from brain and sends it to the muscles.
Ventral columns (sometimes called anterolateral column)- carry pain info up and motor info down.
Brain stem
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Medulla– regulates blood pressure and respiration.
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Ventral pons– pontine nuclei, relay signals from cortex to the cerebellum
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Dorsal pons– respiration taste and sleep
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Midbrain– auditory and visual systems, substantia nigra pars compact (dopaminergic neurons). Deteriorates in Parkinson’s disease.
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Diencephalon
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Contains the thalamus and hypothalamus
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Thalamus– “relay station to the cerebral cortex”- an essential link in the transfer of most sensory information from periphery to cerebral cortex. Also plays a role in filtering information from the periphery.
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Hypothalamus– lies ventral to thalamus. Controls a variety of functions, growth, eating, drinking, maternal behavior by regulating hormonal secretions of the pituitary gland. Connects to virtually every part of brain. Important in initiating and maintaining behaviors that the organism finds rewarding
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The diencephalon contains the…
The thalamus can be generally thought of as the relay station to the cortex.
The hypothalamus lies ventral to the thalamus and controls an array of important physiological functions such as feeding, fluid balance, and hormonal secretions of the endocrine system.
Cerebral Hemispheres
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Largest portion of the human brain
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Cerebral cortex– cognitive functioning
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Hippocampus– memory
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Basal ganglia– control of fine movement
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Amygdala– social behavior and expression of emotion
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Now let’s finally talk about highest order parts of teh central nervous system the cerebral hemispheres.
The two cerebral hemispheres sit atop and surround the diencephalon and much of the brain stem.
Seat of cognition, but it doesn't work alone!
limbic system includes both the amygdala is the integrative center for emotions, emotional behavior, and motivation
Lobes of the cerebral hemispheres
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Primary motor cortex
Primary somatosensory cortex
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Cortical neurons are organized into layers
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Neuronal signaling
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Electrical signals of nerve cells
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Voltage-dependent membrane permeability
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Channels and transporters
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Synaptic transmission
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Neurotransmitters, receptors, and their effects
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Molecular signaling within neurons
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So, how do neurons convey information over long distances that results in information transfer to other neurons at synaptic connections? It through electrical signaling that neurons are able to generate and transmit information. And this electrical signaling is possible because of a combination of…
- voltage-dependent membrane permeability
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- which in turn requires special membrane proteins called ion channels and transporters
- synaptic transmission
- which in turn requires neurotransmitters, their membrane bound protein receptors and their resulting effects
as well as general molecular signaling within neurons as any living cell might have
Types of electrical signals in neurons
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This figure shows these 3 types of neuronal signals.
Here is a receptor potential in a pacinian corpuscle, which is a type of mechanosensory receptor on sensory nerve endings near the surface of your skin.
Here is a synaptic potential recorded in a postsynaptic neuron.
Here is an action potential in a motor neuron. Look as the y-axes here— the action potential has a much larger amplitude change than receptor or synaptic potentials.
Nernst equation
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Statement of the equilibrium condition for a single ion species across a membrane that is permeable only to that ionic species:
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Ex = equilibrium potential in mV
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R = the gas constant (8.3 J mol-1 K-1)
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T = absolute temperature (K)
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F = faraday constant (9.6x104 J mol-1 V-1)
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z = valence of the ion, including sign.
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ln = natural log (base e)
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out concentration of an ion outside; [x]in inside
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RT/F can be a constant at room temperature to give a simplified equation:
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So I stated that the Nernst equation is how we can calculate the equilibrium potential for a cell membrane permeable to one type of ion.
And here is the Nernst equation is:
Where Ex is:
Gas constant R equivalent to the Boltzmann constant but expressed in units of energy. The physical significance of R is work per degree per mole. R = 8.3144598 J mol−1 K−1 == R = work / amount x temperature. Relates the energy scale in physics to the temperature scale, when a mole of particles at the stated temperature is being considered. Joules/mol/K
Faraday constant = magnitude of electric charge per mole of electrons = 96485.33289 C/mol. Expressed in C/mol or J/mol/V
Temperature is:
z is the valence of the ion in question
ln is the natural logarithm which has the mathematical constant e or 2.718 as it’s base.
Now many of the classical experiments recording membrane potential in squid axon or other preparations were conducted at room temperature, which is 20ºC or about 68ºF.
Thus to make calculations simpler in the classic scientific papers (often from the 1930s and 1940s before computers) this equation for experiments carried out at room temperature is often simplified to the following of:
which uses the base10 logarithm. Since —>
ln(x) / log10(x) = 2.30
—> 2.30 * log10(x) = ln(x)
R = 8.3 J/Kmol, T = 37ºC + 273ºC = 310 K, F = 9.610^4 J/mol*V
E =
log(7) / log10(7)
R = 8.3
F = 9.6 * 10^4
T = 20+273
(R*T / F) * 1000 * 2.3
==>58.26427
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K+ concentration gradient determines resting membrane potential
why does it deviate from the line at low K+ concentrations?
increasing extracellular K+ increases resting potential
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So Hodgkin and Katz did this experiment, varying the extracellular K+ concentration while recording the squid axon membrane potential and found that increasing the Kout incr the resting membrane potential.
They plotted resting membrane potential against the extracellular K+ concentration, shown in this red curve.
If internal K+ is unchanged, a plot of membrane potential against the log of external K+ concentration would yield a straight line with slope of 58mV per tenfold change in external K+ concentration at RT.
However it deviates from this expected relationship (shown by the black line), especially at lower K+ concentrations. Why is this?
Because other ions, particularly Cl- and Na+, are also slightly permeable and the contribution of these other ions is more evident at low K+ concentrations.
Role of sodium in the generation of an action potential
- Lowering Na+ decreases both the rate and the rise of an action potential
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When Hodgkin and Katz did this low extracellular Na experiment, the AP had a smaller amplitude and also had a slower or longer timecourse so that the squid axon spiked at slower rate.
The action potential– summary
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Rising phase
Overshoot phase
Falling phase
Undershoot phase
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And this is just a overall summary of what we have been discussing
Electric current flow across a squid axon membrane
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nothing (except for a capacitive transient)
inward and
outward currents
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And so here are the results from this type of voltage clamp experiment.
If you command that the cell membrane potential be hyperpolarized, you get very little or negligible current flowing across the membrane except for a very brief capacitive current that you always see in these voltage clamp experiments.
This is because the cell membrane essentially acts as a parallel RC circuit where a resistor and a capacitor are connected in parallel and to a constant current source. Ion channels are resistors, lipid bilayer with the extracellular and intracellular environments act as capacitor, storing charge in the form of ions accumulating near the surface of the membrane. When a switch is turned on in an RC circuit current flows from the battery to the capacitor until the capacitor is charged to a voltage that is same as the battery.
However when Hodgkin and Huxley depolarized the membrane, a transient inward current occurs followed by a slow outward current.
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A capacitor (originally known as a condenser) is a passive two-terminal electrical component used to store electrical energy temporarily in an electric field. Consists of two parallel conductors. Lipid membrane with the inner and outer cellular environment acts as this. The membrane capacitance per unit areas is mostly constant at about 1 µF/cm2.
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When the voltage is constant, the current through the capacitative pathway is zero because the capacitor has acquired the charge Q (coulombs) according to the relationship Q=CV. C is capacitance (farads) Ic is capacitive current. Ic = C(dV/dt)
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as long as V is changing with time, there will be a current flowing towards the capacitor.
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if V is constant in time, there is no capacitive current.
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product of resistance and capacitance has the unit of time and is called the time constant. Time constant defines how quickly capacitors charge or discharge over time.
http://nerve.bsd.uchicago.edu/med98c.htm
Current produced by membrane depolarizations at several different potentials
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no inward current
outward current
inward current
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This show several different voltage steps (with the brief capacitive current omitted for clarity)
…Notice as we approach ENa the inward current disappears.
Speed of action potential conduction in unmyelinated versus myelinated axons
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Patch clamp
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Allows one to look at currents flowing through a single channel.
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Pipette with small opening makes a tight seal with the membrane.
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Currents are amplified and measured
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Can be adapted to do whole cell recordings, inside out recordings or outside out recordings.
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Measurements of ionic currents flowing through single Na+ channels
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Small inward currents
Open at beginning of pulse
Close quickly
Neuroscience5e Fig. 4.1
Note:
Patch a piece of membrane and block K currents. Do a bunch of short recordings while clamping the membrane at depolarized potential. e.g. here is 7 trials. Notice the amplitude is discrete— it is unitary. If you were recording from lots of…
Transient channel opening in Na+ channels (inward current).
Measurements of ionic currents flowing through single K+ channels
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Early delay in opening
Once open stay open
Neuroscience5e Fig. 4.2
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Sustained channel opening in K+ channels (outward current).
Functional states of voltage-gated Na+ and K+ channels
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Neuroscience5e Fig. 4.3
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Shown here is a model of the functional states for these channels. Notice a few states for Na vs two for K.
11 steps of synaptic transmission
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Neurotransmitter is synthesized and packaged into vesicles.
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An action potential invades the presynaptic terminal.
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Depolarization causes opening of voltage-gated calcium channels.
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There is an influx of Ca2+. 10-4 mM outside 10-7 mM inside. Rushes in fast.
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Calcium causes vesicles to fuse with membrane.
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Neurotransmitter is released into cleft.
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Transmitter binds to receptors on postsynaptic cell
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This opens or closes postsynaptic channels.
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Postsynaptic current flows inside post-synaptic cell.
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Removal of neurotransmitter by glia uptake or enzymatic degradation
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Retrieval of membrane via endocytosis.
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Formal criteria that define a neurotransmitter
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Must be present in the presynaptic neuron.
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Must be released in response to a depolarization and be Ca2+ dependent.
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Must have specific receptors localized on the post-synaptic cell.
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Note: It does not have to function uniquely a neurotransmitter (it may have other functions). e.g. glutamate, glycine, ATP.
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Major categories of neurotransmitters
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Small molecule neurotransmitters– amino acids, purines, biogenic amines.
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Peptide neurotransmitters– 3-36 amino acid polypeptides, often derived from longer polypeptides.
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Acetylcholine
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The neurotransmitter used at the neuromuscular junction. Also used at synapses in visceral motor system and at some CNS synapses– called cholinergic neurons.
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Synthesized from acetyl CoA and choline by choline acetyl transferase (ChAT)– its presence is a good indication that the neuron is cholinergic.
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Removed from synapse by acetylcholine esterase (AChE) has high activity can cleave 5000 molecules per second
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Sarin “nerve gas” is a AChE inhibitor
Note:
ACh: skeletal muscle excitation vs release from vagus nerve that slows down heart beat (cardiac muscle)—
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-Ligand gated channel that depolarizes skeletal muscle fibers vs g-protein coupled receptor that results in hyperpolarization of cardiomyocytes.
Glutamate
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Most important transmitter for normal brain function.
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Nearly all excitatory neurons in the CNS are glutamatergic.
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Does not cross the blood brain barrier.
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Glutamine is most common precursor glutaminase converts it to glutamate.
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Retrieved from synapse by glutamate transporters in glia and neurons. Glia (astrocytes) turn glutamate to glutamine and spit it back out
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Too much glutamate can kill the post-synaptic neuron (excitotoxicity). A major problem after damage due to stroke.
Note:
Most important neurotransmitter for normal brain function. Almost all excitatory neurons in CNS are glutamatergic. Half of all synapses estimated to use this transmitter. Glutamate is non-essential a.a. (by that I mean non-essential per dietary requirements) that does not cross the blood brain barrier. Synthesized inside neurons by local precursors.
histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.
Monosodium glutamate (MSG, also known as sodium glutamate) is the sodium salt of glutamic acid
GABA and glycine
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Most inhibitory neurons use one or the other.
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Inhibits the ability to fire action potentials.
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GABA (gamma-aminobutyric acid) made from glutamate by glutamic acid decarboxylase (GAD), requires Vitamin B6 as cofactor. B6 deficiency can lead to loss of synaptic transmission.
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Glycine– about 1/2 of neurons in spinal cord use glycine.
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Both GABA and glycine are rapidly taken up by glia and neurons.
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Hyperglycinemia– defect in glycine uptake and removal leading to severe mental retardation.
Note:
As many as a third of synapses in the brain use GABA as an inhibitory transmitter. Most commonly found in local circuit neurons.
glycine encephalopathy:
http://ghr.nlm.nih.gov/condition/glycine-encephalopathy
Glycine encephalopathy, which is also known as nonketotic hyperglycinemia or NKH, is a genetic disorder characterized by abnormally high levels of a molecule called glycine. This molecule is an amino acid, which is a building block of proteins. Glycine also acts as a neurotransmitter, which is a chemical messenger that transmits signals in the brain. Glycine encephalopathy is caused by the shortage of an enzyme that normally breaks down glycine in the body. A lack of this enzyme allows excess glycine to build up in tissues and organs, particularly the brain, leading to serious medical problems.
Dopamine
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Produced by the enzyme DOPA decarboxylase
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Made by substantia nigra pars compacta (which connects to corpus striatum for coordination of body movements).
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Does not cross the blood brain barrier, but levadopa (L-DOPA) does.
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Parkinson’s treatments include L-DOPA plus degradation enzyme inhibitors
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Cocaine inhibits uptake of dopamine (inhibits DAT)
Note:
Synthesized in cytoplasm of presynaptic terminals.
Loaded into synaptic vesicles by vesicular monoamine transporter (VMAT). Dopamine in synaptic cleft is terminated by reuptake of dopamine into nerve terminals or glia cells by a Na-dependent dopamine cotransporter called DAT. Cocaine works by inhibiting DAT, increasing dopamine concentrations in synaptic cleft.
Amphetamine also inhibits DAT as well as a transporter for norepinephrine
Catabolized by monoamine oxidase and catechol O-methyltransferase (COMT). Both neurons and glia contain mitochondrial MAO and cytoplasmic COMT. Inhibitors of these enzymes are targets of some kinds of antidepressants (phenelzine and tranylcypromine)
Acts throught GPCRs. D3 parallels that of other metabotropic receptors like mAChR. Subtypes act by activating or inhibiting adenylyl cyclase.
Activation leads to complex behaviors. Antagonists can cause catalepsy (state where difficult to initiate voluntary movement).
-L-DOPA is the precursor to the neurotransmitters dopamine, norepinephrine (noradrenaline), and epinephrine (adrenaline) collectively known as catecholamines.
- -it is converted into dopamine by the enzyme aromatic L-amino acid decarboxylase, also known as DOPA decarboxylase.
Encephalitis lethargica, sleeping sickness, 40 yrs later Oliver Sacks in NYC treats them with L-DOPA
Latinneostriatum
Part of
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Basal ganglia[1]
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Reward system[2][3]
Components
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Ventral striatum[2][3][4
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Dorsal striatum[2][3][4]
The corpus striatum, a macrostructure which contains the striatum, is composed of the entire striatum and the globus pallidus. The lenticular nucleus refers to the putamen together with the globus pallidus.
Neurotransmitter receptors
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Embedded in the plasma membrane of post-synaptic cell.
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Either are ion channels themselves (ionotropic, or ligand-gated ion channel) or interface with ion channels (metabotropic, or G-protein coupled receptors).
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Ultimately, the binding of neurotransmitter causes opening of channels and ion flux. This can lead to depolarization or hyperpolarization of the membrane potential depending on the ion concentrations and the particular ion species flowing in or out.
Note:
Today we will dive a bit deeper into the structure and function of neurotransmitter receptors.
Hypothetical ion channel selectivities and the reversal potential
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So let’s imaging what the current-voltage relationships would look like for different channel selectivities. Remember the reversal potential is when there there is no net ion flux, so it 0 nA on all these graphs and if a channel is selective to only K, it would be equal to the Ek.
If the channel was selective only to Na, than the Erev would be equal to ENa. Same for chloride.
If the channel was a non-selective cation channel (passing both K and Na) than
11Na, 12Mg, 17Cl, 19K, 20Ca
Na+ and K+ movements during EPCs and EPPs
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-90 typical resting potential of a muscle
depolarization
hyperpolarization
nothing
Neuroscience 5e 5.20
Note:
Even though these ionotropic channels opened by ACh are permeable to both Na and K, at the resting membrane potential the EPC is generated primarily by Na influx because of the reduced driving force on K since at Vrest the membrane potential is closer to Ek.
Summation of postsynaptic potentials
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Neuroscience 5e 5.22
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Events from neurotransmitter release to postsynaptic excitation or inhibition
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Glutamate receptors
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NMDA receptor currents require glycine and removal of Mg2+ block
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fig from:
http://www.bris.ac.uk/synaptic/info/glutamate.html
Effector pathways associated with G-protein coupled receptors
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There are many types of alpha, beta, and gamma g-protein subunits allowing a specific and diverse range of downstream responses.
This shows three examples of different heterotrimeric g proteins bound to 3 types of receptors with 3 different cellular responses.
Parallel processing of sensory information
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Totally fascinating to think out how all this works. Talk about which ones we will go over, common principles, all can get linked together.
Compare and contrast sensory systems
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What are the peripheral receptors? What is their receptive field? What neurotransmitters are used?
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How is the information translated into changes in cell potential?
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What are the circuits, how do they get to the cortex?
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What types of perception defects are associated with damage to different components of the system?
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Overall organization of neural structures that control movement
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Lower motor
system
State of muscle
contraction/relaxation
Execute
movement
Output
system
Upper motor
system
Gating
Motor
learning
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Motor pools
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Retrograde labeling of muscles show that the cell bodies of motor neurons are found in ventral horn of the spinal cord.
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Each motor neuron innervates muscle fibers within a single muscle.
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All the motor neurons innervating a single muscle are grouped together in clusters called motor pools.
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Motor pools are located with a slight spread along the A-P axis.
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There is topography along medial-lateral axis of the spinal cord. Neurons that innervate axial musculature (trunk) are located medially, neurons that innervate distal muscles are located laterally.
Note:
Types of motor neurons
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α motor neurons– innervate the extrafusal muscle fibers, the striated muscle fibers that generate the forces needed for movement.
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γ motor neurons– innervate specialized muscle fibers in the muscle spindles that are embedded within connective tissue in the muscle, known as intrafusal muscle fibers. These fibers are also innervated by sensory axons that send info to the brain and spinal cord about the length and tension of muscle.
Note:
Types of motor units
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Slow (S) motor unit– Small motor neurons innervate relatively few muscle fibers and generate small forces. They innervate small “red” muscle fibers that contract slowly but are relatively resistant to fatigue. These are rich in mitochondria and myoglobin, and are important for activities that require sustained muscular contraction such as posture.
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Fast fatigable (FF) motor unit– Large motor neurons innervate larger, more powerful units. Larger α motor neurons innervate larger pale muscle fibers that generate more force, have sparse mitochondria and are easily fatigued.
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Fast fatigue-resistant (FR) motor unit– are of intermediate size, not as fast as FF units but less fatigable.
Note:
Recruitment of motor neurons to medial gastrocnemius (leg muscle)
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slow for standing
FR for walking or running
FF for sprinting, jumping
Comparison of the function of muscle spindles and Golgi tendon organs
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Comparison of the function of muscle spindles and Golgi tendon organs
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Overview of descending motor control
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Neuroscience 5e Fig. 17.1
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med ventral horn has lower motor neurosn for posteure balance and orienting movements of head and neck during shits of visual gaze. receipve descending input from the pathways orginating mainly in the brainstem, course through the anterior medial white matter of the spional cord and terminate bilaterally.
lateral ventral horn contains lower motor neurons that mediate skilled voluntary movements of the distal extremities. Receive descending projection from the contralateral motor cortex via lateral division of the corticospinal tract.
The medial descending motor pathways
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Overview of descending motor control
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somatotopic organization of the ventral horn in the cervical enlargement. Locations of descending projections from the motor cortex in the lateral white matter and from the brainstem in the anterior-medial white matter are shown.
Section of pyramidal tracts in monkeys produces loss of independent digit control
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Intact (normal)
After section of
corticospinal fibers
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corticalspinal, lateral dorsal input for control of distal/fine movements of the fingers.
Modulation of movement by the basal ganglia
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Basal ganglia do not project directly to the spinal cord, instead they influence movements by regulating the activity of upper motor neurons.
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The basal ganglia are a large set of nuclei that lie deep within the cerebral hemispheres.
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Three main nuclei– caudate, putamen, and the globus pallidus.
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Together with the substantia nigra and the subthalamic nucleus make a loop that links most areas of the cortex with upper motor neurons.
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These neurons are required for the normal course of voluntary movements. Supervise motor movements.
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Direct and indirect pathways through the basal ganglia
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Dopamine excites the direct and inhibits the indirect pathway
Neuroscience 5e Fig. 18.7
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Hemiballismus: violent involuntary movements of the limbs
- Defects in the subthalamic nucleus of the contralateral side of the movements
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Summary explanation of hypokinetic and hyperkinetic disorders
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Summary explanation of hypokinetic and hyperkinetic disorders
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Descending systems that control somatic and visceral motor pathways in the expression of emotion
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Neuroscience 5e Fig. 29.2
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Pathways involved in fear conditioning
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Neuroscience 5e Fig. 29.5
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Functional and anatomical organization of the limbic loop through the basal ganglia
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Nucleus accumbens– contains MSNs
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Ventral tegmental area (VTA)– releases dopamine
Nucleus accumbens
Neuroscience 5e Fig. 29.10
Note:
Much like the direct pathway. Inputs from different parts of cortex, including amygdala.
to MSNs in ventral striatum the nucleus accumbens. These gabaergic projections then inhibit inhibitory projections in the in the ventral globus pallidus called the ventral pallidum. So there is a disinhibitory effect, much as we discussed before for other basal ganglia loops.