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Hearing the call of Neurons Winter 2017 Peter Woodruff Shining a light on the retina Problems: 1. Tissues are . How do we see the details inside? 2. Most cells are more or less How do we tell one from another? 3. Neurons are often long and thin. How do we follow the branches? Left: schematic drawing by Santiago Ramon y Cajal (circa 1900). Right: section through a rat retina. Neurons and Synapses Microscopes first used in 17th century For 200 years, nervous system seen largely as fatty globules in tissue Then: • Discovery of neurons with their dendrites and axons • Neuron doctrine • Discovery of synapses and chemical transmission What led to these changes? Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization Microscope 2. Thin slicing 3. Staining to distinguish cell / tissue parts 1. Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. Microscope ~1670 Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. Microscope x-section of optic nerve: Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. Microscope Cork “cells” through microscope Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. Microscope Thin slicing How to cut soft, squishy tissue? Why cut? Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. Microscope Thin slicing How to cut soft, squishy tissue? Why cut? Fresh tissue sample Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. Microscope Thin slicing How to cut soft, squishy tissue? Why cut? Ideally: use a sharp knife, hard material Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. Microscope Thin slicing How to cut soft, squishy tissue? Why cut? Ideally: use a sharp knife, hard material So: harden tissue! How? Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. Microscope Thin slicing How to cut soft, squishy tissue? Why cut? Ideally: sharp knife, hard material So: harden tissue! How? Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. Microscope Thin slicing How to cut soft, squishy tissue? Why cut? Ideally: sharp knife, hard material So: harden tissue! How? ? Or replace water with wax or plastic. Take sample, stop enzymes, remove water, replace with wax or plastic Take sample, stop enzymes, remove water, replace with wax or plastic Now we need to cut! Very carefully! Cutting the sample: use a microtome Cutting the sample Cutting the sample A ribbon of 4 mm sections being cut from a paraffin block using a rotary microtome. Next: flatten and put onto microscope slide. The “Big Brain” project cutting it close. A special microtome cuts a human brain preserved in paraffin wax into 20-micrometre thick slivers to help map its anatomical structure with high resolution Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 1. 2. 3. Microscope Thin slicing Staining to distinguish cell / tissue parts Colored dyes stick to some parts more than others Stained slides cerebellum Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 4. Cell Theory: Schleiden and Schwann + ~1839: • All living organisms are composed of one or more cells. BUT viruses? • The cell is the basic unit of structure and organization in organisms. • Cells arise from pre-existing cells. Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 5. Neurons are cells (1890s) with dendrites, cell bodies and axons Retinal neurons have many different shapes and sizes. A midget bipolar and a parasol-type ganglion cell are shown. Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 5. Neurons are cells (1890s) with dendrites, cell bodies and axons Neurons and Synapses Discoveries depended on improvements in instruments, experimental technique and conceptualization 6. Camillo Golgi developed silver staining technique which only stained some neurons (defective?) 7. Santiago Ramon Y Cajal improved Golgi’s technique Santiago Ramón y Cajal and the Neuron Doctrine Santiago Ramón y Cajal, from the book “The Beautiful Brain.” A diagram suggesting how the eyes might transmit a unified picture of the world to the brain; a purkinje neuron from the human cerebellum, circa 1900; and a diagram showing the flow of information through the hippocampus in the brain Ramón y Cajal A self-portrait of Ramón y Cajal in his laboratory in Valencia, Spain, about 1885 artist, photographer, doctor, bodybuilder, scientist, chess player and publisher. He was also the father of modern neuroscience. What he saw Pyramidal cells stained with the Golgi method improved by Ramón y Cajal. This method, developed by the Italian scientist Camillo Golgi, made it possible to see the details of a whole neuron without the interference of its neighbors. In 1906 he and Golgi shared a Nobel Prize. Large pyramidal neuron in the cerebral cortex, the outside part of the brain, visible by naked eye (as a dot) Purkinje neuron from cerebellum, illustrating its treelike structure in great detail Neuron Types The nervous system has many types of cells, not just neurons Neuron Doctrine 1. The neuron is the structural and functional unit of the nervous system 2. Neurons are individual cells not continuous with other cells 3. Neurons have three functional parts: dendrites, soma and axon. 4. Conduction preferentially flows from dendrites through soma to ends of the axon (axon terminals, with their synapses) Nerve impulse passing along axon dendrites axon • Unidirectional • All-or-nothing • Delay after firing Message flows down axon to synapse. Transmission to post-synaptic cell affects that cell’s behavior: may become more easily excited (excitatory synapse) or may become less easily excited (inhibitory synapse) Neuron Doctrine 5. Synapse identified (1846), could be electrical or chemical transmission Unidirectional (generally axon to dendrite) Excitatory and inhibitory action Delay in transmission Neuron Doctrine Santiago Ramón y Cajal and the Neuron Doctrine Hunched Over a Microscope, He Sketched the Secrets of How the Brain Works By JoAnna Klein, New York Times https://www.nytimes.com/2017/02/17/science/santiago-ramon-y-cajal-beautiful-brain.html?_r=0 The Beautiful Brain: The Drawings of Santiago Ramón y Cajal Saturday, Jan 28 2017 - Sunday, May 21 2017 Minneapolis; Vancouver, British Columbia; New York; Cambridge, Mass.; and Chapel Hill, N.C. How can one sense movement of a fluid? How to sense movement of a fluid? Hair versus hair cells How to sense movement of a fluid? How to sense movement of a fluid? Sensitive Sharks Sensitive Sharks Linear Acceleration Coding by Maculae Tilt causes shearing forces on some hair cells which depolarize as others hyperpolarize. Rotational Coding by Semicircular Canals As one of the canals moves in an arc with the head, the internal fluid moves in the opposite direction, causing the cupula and stereocilia to bend. Brain interprets relative activation of all six canals to give precise indication of head movement. Sound attributes • Frequency: vibrations per second, in hertz, Hz • Amplitude: sound pressure a sound intensity, as in decibels, dB • Temporal variation: eg: duration • Tone: simple sound • Complex sound: contains >1 frequency Fun Facts about Hair Cells • Hair cells can detect deflections of 0.3 nanometers (< size of atom!) • Threshold of hearing: 1 billionth of atmospheric pressure • Can convert stimulation into nerve impulse in 10 microseconds • Threshold of pain at 130 dB is 1013 times more intense than threshold of hearing, at 0 dB! When the stereocilia are bent in response to a sound wave, an electromotile response occurs. With every sound wave, the cell shortens and then elongates. This pushes against the tectoral membrane, selectively amplifying the vibration of the basilar membrane.