BioPsy- Methods and Strategies of Research PDF

METHODS AND STRATEGIES OF RESEARCH STEREOTAXIC SURGERY “Stereotaxis” – solid arrangement Experimental Ablation – the removal or Brain surgery using a stereotaxic apparatus to destruc...

METHODS AND STRATEGIES OF RESEARCH STEREOTAXIC SURGERY “Stereotaxis” – solid arrangement Experimental Ablation – the removal or Brain surgery using a stereotaxic apparatus to destruction of a portion of the brain of a position an electrode or cannula in a specified laboratory animal position of the brain. - Latin “ablatus” – “carrying away” Stereotaxic Atlas Lesion study – an experimental study in The heads of newborn babies contain a soft which the behavior of animals with brain spot at the junction of the coronal and sagittal lesions is studied sutures called fontanelle. - To discover what functions are performed by Once this gap closes, the junction is called different regions of the brain and then to bregma. understand how these functions are combined Bregma -a junction of the sagittal and coronal to accomplish particular behaviors. sutures of the skull Often used as a reference point for stereotaxic PRODUCING BRAIN LESIONS brain surgery Anesthetize the animal, cut its scalp, remove Stereotaxic atlas – a collection of drawings of part of its skull, and cut through the dura mater, sections of the brain of a particular animal bringing the cortex into view. Then we can use a with measurements that provide coordinates suction device to aspirate the brain tissue. To for stereotaxic surgery. accomplish this tissue removal, we place a glass pipette on the surface of the brain and suck away brain Stereotaxic Apparatus tissue with a vacuum pump attached to the pipette by a A device that permits a surgeon or cannula flexible tube. into a specific part of the brain. In destroying regions that are hidden away in the depths of the brain, brain lesions of subcortical HISTOLOGICAL METHODS regions are usually produced by passing electrical Fixation current through a stainless steel wire that is coated Sectioning with an insulating varnish except for the very tip. Staining Electron microscopy RF lesion - a brain lesion produced by passing radio FIXATION AND SECTIONING frequency (RF) current through a fine wire inserted into the brain. Fixative – a chemical such as formalin - Lesions produced by these means destroy - Used to prepare and preserve body tissue everything in the vicinity of the electrode tip. Formalin – the aqueous solution of formaldehyde gas - The most commonly used tissue fixative Excitotoxic lesion - a brain lesion produced by intracerebral It halts autolysis, hardens the very soft and injection of an excitatory amino acids fragile brain, and kills any microorganisms that might - A more selective method destroy it. - The selectivity of excitotoxic lesions permits the investigator to determine whether the Microtome – an instrument that produces very behavioral effects of destroying particular thin slices of body tissues. brain structure are caused by the death of neurons located there or by the destruction of Slices prepared for examination under a light axons that pass nearby. microscope are typically 10 to 80 μm in thickness; those prepared for the electron microscope are Sham lesion generally cut at less than 1 μm (A μm, or micrometer, - a “placebo” procedure that duplicates all the is one-millionth of a meter, or one-thousandth of a steps of producing a brain lesion except for the millimeter. one that actually causes the brain damage. Slices of brain tissue is called sections STAINING Fluorogold A dye that serves as a retrograde label Franz Nssl – discovered that a dye known as Taken up by terminal button and carried back methylene blue would stain the cell bodies of brain to the cell bodies tissue. The material that takes up the dye, known as STUDY OF THE LIVING HUMAN BRAIN the Nssl substance, consists of RNA, DNA, and associated proteins located in the nucleus Computerized tomography (CT) and scattered, in the form of granules in the the use of a device to analyze data obtained by cytoplasm. a scanning beam of X-rays to produce a two-dimensional picture of a slice through the Cresyl violet – most frequently used stain body. uses dyes such as cresyl violet and other Nissl dyes to estimate the number of cell bodies in a particular part of the brain by counting the Nissl stained dots. ELECTRON MICROSCOPY Scanning electron microscope – a microscope that provides three-dimensional information about the Magnetic RResonance Imaging (MRI) shape of the surface of a small object. A technique whereby the interior of the body used to get information about the detailed can be accurately images structure of neurons. uses a scanning electron microscope to obtain an electron micrograph. However, due to a detailed picture of the neurons, it becomes difficult to visualize general aspects of neuron structure. TRACING NEURAL CONNECTIONS TRACING EFFERENT AXONS Anterograde labeling method A histological method that labels the axons and terminal buttons of neurons whose cell bodies are located in a particular region. PHA-L - Phaseolus vulgaris leukoagglutinin A protein derived from kidney beans used as RECORDING AND STIMULATING NEURAL an anterograde tracer ACTIVITY, NEUROCHEMICAL METHODS, Taken up by dendrites and cell bodies and GENETIC METHODS carried to the ends of the axons Recording Neural Activity TRACING AFFERENT AXONS Axons produce action potentials, and terminal buttons elicit postsynaptic potentials in the Retrograde labelling method membrane of the cells with which they form a histological method that labels cell bodies synapses. that give rise to the terminal buttons that form Recording can be made chronically, over an synapses with cells in a particular region. extended period of time after an animal Moving backward recovers from surgery, or acutely, for a relatively short period of time during which an animal is kept anesthetized. RECORDING WITH MICROELECTRODES STIMULATING NEURAL ACTIVITY Electrical or chemical stimulation Microelectrode Electrical stimulation involves passing an a very fine electrode, generally used to record electrical current through a wire inserted into activity of individual neurons. the brain. Single-unit recording Chemical stimulation is accomplished by recording the electrical activity of a single injecting a small amount of an excitatory neuron. amino acids into the brain ○ Disadvantage: slightly more RECORDING WITH MACROELECTRODES complicated, as it requires cannulas, tubes, special pumps or syringes, and a Macroelectrode terile solutions of excitatory amino an electrode used to record the electrical acids. activity of large numbers of neurons in a ○ Advantage: it activates cell bodies but particular region of the brain; much larger than not axons. Thus, the effects of microelectrode chemical stimulation are more Electroencephalogram(ECG) localized than the effects of electrical an electrical brain potential recorded by stimulation. placing electrodes on the scalp. NEUROCHEMICAL METHODS RECORDING THE BRAIN’S METABOLIC AND SYNAPTIC ACTIVITY Finding Neurons that produce Particular Neurochemicals 2-deosyglucose (2-DG) a sugar that enters cells along with glucose but Immunocytochemical method is not metabolized. a histological method that uses radioactive Autoradiography antibodies or antibodies bound with a dye a procedure that locates radioactive substance molecule to indicate the presence of particular in a slice of tissue proteins or peptides. The radiation exposes a photographic Localizing Particular Receptors emulsion or a piece of film that covers the To identify the neurons that receive input, an tissue experimenter use an anterograde labeling Fos method to trace the pathway of axons a protein produced in the nucleus of the neuron in response to synaptic stimulation. GENETIC METHODS Positron emission tomography(PET) Twin Studies The use of a device that reveals the Adoption Studies localization of a radioactive tracer in a living brain. Targeted mutation Functional MRI (fMRI) mutated genes that reproduced in the a modification of the MRI procedure that laboratory and inserted into the chromosomes permits the measurement of regional of mice. metabolism in the brain. MEASURING THE BRAIN’S SECRETION Microdialysis a procedure for analyzing chemicals present in the interstitial fluid through a small piece of tubing made of a semipermeable membrane that is implanted in the brain. High-performance liquid chromatography