Biopsychology Research Methods PDF

Chapter 5 The Research Methods of Biopsychology Understanding What Biopsychologists Do The Photolibrary Wales/Alamy Stock Photo Chapter Overview and Learning Objectives PART ONE Methods of...

Chapter 5 The Research Methods of Biopsychology Understanding What Biopsychologists Do The Photolibrary Wales/Alamy Stock Photo Chapter Overview and Learning Objectives PART ONE Methods of Studying the Nervous System Methods of Visualizing LO 5.1 Describe two x-ray-based techniques for visualizing the living and Stimulating the Living human brain. Human Brain LO 5.2 Describe the positron emission tomography (PET) technique. LO 5.3 Describe three magnetic-field-based techniques for imaging the living human brain. LO 5.4 Describe an ultrasound-based technique for imaging the living human brain. LO 5.5 Describe three transcranial stimulation techniques. Recording Human LO 5.6 Describe two psychophysiological measures of brain activity. Psychophysiological LO 5.7 Describe two psychophysiological measures of somatic Activity nervous system activity. 121 M05_PINE1933_11_GE_C05.indd 121 22/01/2021 10:51  122 Chapter 5 LO 5.8 Describe two psychophysiological measures of autonomic ­nervous system activity. Invasive Physiological LO 5.9 Describe the process of stereotaxic surgery. Research Methods LO 5.10 Describe four types of lesion methods and explain why it is important to be cautious when interpreting the effects of lesions. LO 5.11 Describe the technique of electrical brain stimulation. LO 5.12 Describe four invasive electrophysiological recording methods. Pharmacological Research LO 5.13 Describe the various methods of drug administration. Methods LO 5.14 Describe the method of selective neurotoxic lesions. LO 5.15 Describe two techniques for measuring chemical activity in the brain. LO 5.16 Describe two techniques for locating particular ­neurotransmitters or receptors in the brain. Genetic Methods LO 5.17 Explain the gene knockout technique by describing an ­experiment that employed the technique. LO 5.18 Explain the gene knockin technique by describing an experiment that employed the technique. LO 5.19 Describe how modern gene-editing techniques, such as the CRISP-Cas9 method, can provide better ways of assessing the role of a gene in behavior. LO 5.20 Explain how green fluorescent protein has been used as a research tool in the neurosciences. LO 5.21 Explain how opsins have been used as a research tool in the neurosciences. PART TWO Behavioral Research Methods of Biopsychology Neuropsychological Testing LO 5.22 Describe three approaches to neuropsychological testing. LO 5.23 Describe those tests that are often administered as part of an ­initial common neuropsychological test battery. LO 5.24 Describe tests that might be used by a neuropsychologist to investigate in more depth general problems revealed by a ­common neuropsychological test battery. Behavioral Methods of LO 5.25 Describe the paired-image subtraction technique. Cognitive Neuroscience LO 5.26 Understand the default mode network and know the structures that are part of that network. LO 5.27 Explain what a mean difference image is. LO 5.28 Explain the concept of functional connectivity. M05_PINE1933_11_GE_C05.indd 122 22/01/2021 10:51  The Research Methods of Biopsychology 123 Biopsychological LO 5.29 Describe three behavioral paradigms used to study Paradigms of Animal ­species-­common behaviors. Behavior LO 5.30 Describe the Pavlovian conditioning paradigm and the operant conditioning paradigm. LO 5.31 Describe four seminatural animal learning paradigms. LO 5.32 Explain why multiple techniques should be used when trying to answer a specific question. As Professor P. was driving home from his pre-surgery The Ironic Case of Professor P. tests, his mind wandered from his current plight to his day at the hospital. “Quite interesting,” he thought to himself. There were Two weeks before his brain surgery, Professor P. reported to the biopsychologists everywhere, doing biopsychological things. In hospital for a series of tests. What amazed Professor P. most all three labs he had visited, there were people who had begun about these tests was how familiar they seemed. No, Professor P. their training as biopsychologists. was not a psychic; he was a biopsychologist, and he was struck Two weeks later, Professor P. was rolled into the prepara- by how similar the tests performed on him were to the tests he tion room. “Sorry to do this, Professor P., you were my favorite had encountered in his work. instructor,” the nurse said, as she inserted a large needle into Professor P. had a brain tumor on his right auditory-vestibular Professor P.’s face and left it there. cranial nerve (cranial nerve VIII; see Appendices III and IV), and he Professor P. didn’t mind; he was barely conscious. He did had to have it excised (cut out). First, Professor P.’s auditory abilities not know that he wouldn’t regain consciousness for several were assessed by measuring his ability to detect sounds of various days—at which point he would be incapable of talking, eating, volumes and pitches and then by measuring the magnitude of the or even breathing. But more about that later. EEG signals evoked in his auditory cortex by clicks in his right ear. Next, Professor P.’s vestibular function (balance) was tested by injecting cold water into his ear. Don’t forget Professor P.; you will learn more about him in “Do you feel anything, Professor P.?” Chapter 10. For the time being, this case demonstrates that “Well, a cold ear.” many of the fundamental research methods of biopsychol- “Nothing else?” ogy are also used in clinical settings. Let’s move on to the “No.” methods themselves. So colder and colder water was tried with no effect until the final, coldest test was conducted. “Ah, that feels weird,” said Professor P. “It’s kind of like the bed is tipping.” The results of the tests were bad, or good, depending on PART ONE Methods of your perspective—they certainly revealed his deficits. Professor Studying the Nervous System P.’s hearing in his right ear was poor, and his right vestibular This is the first of the two parts that compose this chapter. nerve was barely functioning. “At the temperatures we flushed In this part, we present the methods used by biopsycholo- down there, most people would have been on their hands gists to study the nervous system. As you will soon see, the and knees puking their guts out,” said the medical technician. ­Professor P. smiled at her technical terminology. methods are extremely diverse. Of course, he was upset that his brain had deteriorated so badly, but he sensed that his neurosurgeon was secretly pleased: “We won’t have to try to save the nerve; we’ll just cut it.” Methods of Visualizing There was one last test. The skin of his right cheek was lightly pricked while the EEG responses of his somatosensory cor- and Stimulating the Living tex were recorded from his scalp. “This is just to establish a base- line for the surgery,” it was explained. “One main risk of removing Human Brain tumors on the auditory-vestibular cranial nerve (VIII) is damaging This module first describes four different sorts of methods the facial cranial nerve (VII), and that would make the right side of your face sag. So during the surgery, electrodes will be inserted for visualizing the living human brain: x-ray-based tech- in your cheek, and your cheek will be repeatedly stimulated with niques, a radioactivity-based technique, magnetic-field- tiny electrical pulses. The cortical responses will be recorded and based techniques, and an ultrasound-based technique. fed into a loudspeaker so that the surgeon can immediately hear Next, it presents three techniques for noninvasively stimu- changes in the activity if his scalpel starts to stray into the area.” lating the living human brain. M05_PINE1933_11_GE_C05.indd 123 22/01/2021 10:51  124 Chapter 5 substance then heightens the contrast between the compart- Figure 5.1 A cerebral angiogram of a healthy human. ment and the ­surrounding tissue during x-ray photography. One contrast x-ray technique, cerebral angiography, uses the infusion of a radio-opaque dye into a cerebral artery to visualize the cerebral circulatory system during x-ray photography (see Figure 5.1). Cerebral angiograms are most useful for localizing vascular damage, but the dis- placement of blood vessels from their normal position also can indicate the location of a tumor. Journal Prompt 5.1 Egas Moniz, the inventor of the lobotomy, was also the pioneer of cerebral angiography. Some have argued that Moniz’s Nobel Prize for the lobotomy should have been revoked. However, others have argued that he would have won it anyway for his important work on cerebral CNRI/Science Source angiography. Do you think Moniz deserved to win the Nobel Prize? Why or why not? X-Ray-Based Techniques COMPUTED TOMOGRAPHY. In the early 1970s, the LO 5.1 Describe two x-ray-based techniques for introduction of computed tomography revolutionized the visualizing the living human brain. study of the living human brain. Computed tomography (CT) is a computer-assisted x-ray procedure that can be Prior to the early 1970s, biopsychological research was impeded used to visualize the brain and other internal structures of by the inability to obtain images of the organ of primary inter-the living body. During cerebral computed tomography, the est: the living human brain. Conventional x-ray photography neurological patient lies with his or her head positioned is useless for this purpose. When an x-ray photograph is taken, in the center of a large cylinder, as depicted in Figure 5.2. an x-ray beam is passed through an object and then onto a photographic Figure 5.2 Computed tomography (CT) uses x-rays to create a brain scan. plate. Each molecule the beam has passed through absorbs some of the X-ray source X-ray detector Horizontal CT scans radiation; thus, only the unabsorbed portions of the beam reach the photo- graphic plate. This makes x-ray pho- tography effective in characterizing internal structures that absorb x-rays differently than their ­surroundings— just like a revolver in a suitcase full of clothes or a bone surrounded by flesh. However, by the time an x-ray beam has passed through the numer- ous overlapping structures of the brain, which differ only slightly in their ability to absorb x-rays, it carries little information about the structures through which it has passed. Three-dimensional reconstruction CONTRAST X-RAYS. Although conventional x-ray photography is not useful for visualizing the brain, contrast x-ray techniques are. ­Contrast x-ray techniques involve injecting into one compartment of the body a substance that absorbs x-rays either less than or more than the surrounding tissue. The injected M05_PINE1933_11_GE_C05.indd 124 22/01/2021 10:51  The Research Methods of Biopsychology 125 On one side of the cylinder is an x-ray tube that proj- The most significant current application of PET ects an x-ray beam through the head to an x-ray detector t­echnology is its use in identifying the distribution of par- mounted on the other side. The x-ray tube and detector ticular molecules (e.g., neurotransmitters, receptors, trans- rotate rapidly around the head of the patient at one level porters) in the brain (see Camardese et al., 2014). This is of the brain, taking many individual x-ray photographs accomplished by injecting volunteers with radioactively as they rotate. The meager information in each x-ray labeled ligands (ions or molecules that bind to other mol- ­photograph is combined by a computer to generate a CT ecules). Then, PET scans can document the distribution of scan of one horizontal section of the brain. Then the x-ray radioactivity in the brain. tube and detector are moved along the axis of the patient’s body to another level of the brain, and the process is repeated. Scans of eight or nine horizontal brain sections Magnetic-Field-Based Techniques are typically obtained from a patient. When combined, LO 5.3 Describe three magnetic-field-based techniques these images provide three-dimensional representations for imaging the living human brain. of the brain. MAGNETIC RESONANCE IMAGING. Magnetic ­resonance Radioactivity-Based Techniques imaging (MRI) is a structural brain-imaging procedure in which high-resolution images are constructed from the LO 5.2 Describe the positron emission tomography measurement of radio-frequency waves that hydrogen (PET) technique. atoms emit as they align with a powerful magnetic field. Positron emission tomography (PET) was the first brain- Such imaging is possible because: (1) water contains two imaging technique to provide images of brain activity hydrogen atoms (H2O) and (2) different brain structures (functional brain images) rather than images of brain struc- contain different amounts of water. This, in turn, means ture (structural brain images). In one common version of that the number of hydrogen atoms differs between brain PET, radioactive fluorodeoxyglucose (FDG) is injected structures, and, therefore, the radio-frequency waves emit- into the patient’s carotid artery (an artery of the neck that ted by a particular brain structure will be different from feeds the ipsilateral cerebral hemisphere). Because of its its neighboring brain structures. MRI provides clearer similarity to glucose, the primary metabolic fuel of the images of the brain than does CT (see Lerch et al., 2017). brain, fluorodeoxyglucose is rapidly taken up by active A two-dimensional MRI scan of the midsagittal plane of the (energy-consuming) cells. However, unlike glucose, brain is presented in Figure 5.4. fluorodeoxyglucose cannot be metabolized; it therefore In addition to providing relatively high spatial accumulates in active neurons and astrocytes until it is ­resolution (the ability to detect and represent differences gradually broken down (see Zimmer et al., 2017). Each in spatial location), MRI can produce images in three PET scan is an image of the lev- els of radioactivity (indicated by Figure 5.3 A pair of PET scans. A scan was done when the volunteer’s eyes were color coding) in various parts of either open (left) or closed (right). Areas of high activity are indicated by reds and ­yellows. one horizontal level of the brain. Notice the high level of activity in the visual cortex of the occipital lobe when Thus, if a PET scan is taken of a the ­volunteer’s eyes were open. patient who engages in an activ- ity such as reading for about 30 seconds after the FDG injec- tion, the resulting scan will indi- cate the areas of the target brain level that were most active during the 30 seconds (see Figure 5.3). Notice from Figure 5.3 that PET scans are not really images of the brain. Each PET scan is merely a colored map of the amount of radioactivity in each of the tiny cubic voxels (volume pixels) that compose the scan. Exactly how each voxel maps onto a particular brain structure can be estimated only by superimposing the scan Occipital lobe on a brain image. NIH/Science Source M05_PINE1933_11_GE_C05.indd 125 22/01/2021 10:51  126 Chapter 5 these new MRI techniques has been diffusion tensor MRI. Figure 5.4 A color-enhanced midsagittal MRI scan. Diffusion tensor MRI is a method of identifying those pathways along which water molecules rapidly diffuse (see Jbadi et al., 2015). Because tracts (bundles of axons) are the major routes of rapid water diffusion in the brain, diffu- sion tensor imaging provides an image of major tracts—see Figure 5.6. Most brain research focuses on the structures of the brain. However, in order to understand how the brain works, it is imperative to understand the connections among those structures—the so-called connectome (see Park & Friston, 2013; Glasser et al., 2016; Swanson & Lichtman, 2016). This is why diffusion tensor images have become a focus of neuroscientific research. Complete descriptions of connectomes already exist for some organisms, including the nematode C. elegans and the mouse (see Oh et al., 2014). Work on the so-called Human Connectome Project is well underway. FUNCTIONAL MRI. MRI technology has been used to produce functional images of the brain. Indeed, functional MRI has become the most influential tool of cognitive ­neuroscience. It is often used to determine if a brain is dys- Scott Camazine/Science Source functional, but it is also used for a variety of other purposes; for example, to infer the content of an individual’s dreams (see Horikawa et al., 2013; Underwood, 2013). dimensions. Figure 5.5 shows a three-dimensional MRI scan Functional MRI (fMRI) produces images repre- of a patient with a growing tumor. senting the increase in oxygenated blood flow to active DIFFUSION TENSOR MRI. Many variations of MRI have areas of the brain. Functional MRI is possible because of been developed. Arguably, one of the most innovative of two attributes of oxygenated blood. First, active areas of the brain take up more oxygenated blood than they need for their energy requirements, and thus oxygen- Figure 5.5 MRI of a growing tumor. The tumor is ated blood accumulates in active areas of the brain (see colored red. Figure 5.6 Diffusion tensor MRI. This three-dimensional image shows the major tracts of the brain. Simon Fraser/Science Source Image Source/Alamy Stock Photo M05_PINE1933_11_GE_C05.indd 126 22/01/2021 10:52  The Research Methods of Biopsychology 127 Hillman, 2014). Second, oxygenated blood has different many neural responses, such as action potentials, occur in magnetic properties than does deoxygenated blood, and the millisecond range. this ­difference influences the radio-frequency waves emit- ted by hydrogen atoms in an MRI. The signal recorded by fMRI is called the BOLD signal (the blood-oxygen-level- Ultrasound-Based Techniques dependent signal). The BOLD signal indicates the parts of LO 5.4 Describe an ultrasound-based technique for the brain that are active or inactive during a cognitive or imaging the living human brain. behavioral test, and thus it suggests the types of analyses Functional ultrasound imaging (fUS) is a new imaging the brain is performing. Because the BOLD signal is the technique that uses ultrasound (sound waves of a higher result of blood flow through the brain, it is important to frequency than we can hear) to measure changes in blood remember that it is not directly measuring the electrical volume in particular brain regions. When a brain region activity of the brain. becomes active, blood levels increase there, and alter the Functional MRI has three advantages over PET: (1) passage of ultrasound through that brain region. nothing has to be injected into the volunteer; (2) it provides As a functional brain imaging method, fUS has three both structural and functional information in the same key advantages over PET and fMRI: (1) it is cheap, (2) highly image; and (3) its spatial resolution is better. A functional portable; and (3) can be used for imaging some individuals, MRI is shown in Figure 5.7. such as human infants, who cannot undergo PET or fMRI It is important not to be unduly swayed by the impres- (see Deffieux et al., 2018). siveness of fMRI images and technology. The images are often presented—particularly in the popular press or gen- eral textbooks—as if they are actual pictures of human Transcranial Stimulation neural activity. They aren’t: They are images of the BOLD LO 5.5 Describe three transcranial stimulation signal, and the relation between the BOLD signal and neu- techniques. ral activity is complex (see Hillman, 2014). Furthermore, fMRI technology has poor temporal resolution, that is, it PET, fMRI, and fUS have allowed cognitive neuroscien- is poor at specifying the timing of neural events. Indeed, tists to create images of brain activity while volunteers it takes 2 or 3 seconds to measure the BOLD signal, and are engaging in particular cognitive activities. Although technically impressive, these kinds of studies of brain activity and cognition all have the same shortcoming: They can be used to show a correlation between brain Figure 5.7 Functional magnetic resonance image (fMRI). activity and cognitive activity, but they can’t prove that This image illustrates the areas of cortex that became more the brain activity caused the cognitive activity (Sack, active when the volunteers observed strings of letters and 2006). For example, a brain-imaging technique may show were required to specify which strings were words—in the control condition, volunteers viewed strings of asterisks that the cingulate cortex becomes active when volunteers (Kiehl et al., 1999). This fMRI illustrates surface activity; but view disturbing photographs, but it can’t prove that the images of sections through the brain can also be displayed. cingulate activity causes the emotional experience—there are many other explanations. There are two obvious ways of supporting the hypothesis that the cingulate cortex is an area for emotional experience. One way would be to assess emotional experience in people lacking a func- tional cingulate cortex. This can be accomplished by studying patients with cingulate damage or by “turning off” the cingulate cortex of healthy patients—transcranial magnetic stimulation is a way of turning off particular areas of cortex. A second way would be to assess emo- tional experiences of volunteers after “turning on” their cingulate cortex—transcranial electrical stimulation and transcranial ultrasound stimulation are ways of turning on areas of cortex. Let us briefly introduce you to transcranial magnetic stimulation and transcranial electrical stimulation, which are currently playing a major role in establishing the causal effects of human cortical activity on cognition and Kent Kiehl/Peter Liddle/University of British Columbia Department of Psychiatry behavior. Transcranial magnetic stimulation (TMS) is a M05_PINE1933_11_GE_C05.indd 127 22/01/2021 10:52  128 Chapter 5 technique that can be used to turn off an area of human cortex by creating a magnetic field under a coil positioned next to the skull (e.g., Candidi et al., 2015). The magnetic Recording Human stimulation temporarily turns off part of the brain while the effects of the disruption on cognition and behavior are Psychophysiological assessed. Although there are still fundamental questions about safety, depth of effect, and mechanisms of neural Activity disruption (see Polanía, Nitsche, & Ruff, 2018; Romei, Thut, The preceding module introduced you to structural & Silvanto, 2016), TMS is often employed to circumvent and functional brain imaging. This module deals with the difficulty that brain-imaging studies have in deter- ­psychophysiological recording methods (methods of record- mining causation. Using different stimulation parameters, ing physiological activity from the surface of the human TMS can also be used to “turn on” an area of cortex body). Six of the most widely studied psychophysiologi- (see Rossini et al., 2015). cal measures are described: two measures of brain activ- Transcranial electrical stimulation (tES) is a technique ity (the scalp EEG and magnetoencephalography), two that can be used to stimulate (“turn on”) an area of the measures of somatic nervous system activity (muscle cortex by applying an electrical current through two elec- ­tension and eye movement), and two measures of auto- trodes placed directly on the scalp. The electrical stimula- nomic nervous system activity (skin conductance and tion temporarily increases activity in part of the brain while ­cardiovascular activity). the effects of the stimulation on cognition and behavior are assessed (see Polanía, Nitsche, & Ruff, 2018). Psychophysiological Measures of The use of tES for its putative cognitive enhancement effects has become popular, and there are many relatively Brain Activity inexpensive tES systems available for purchase online (see LO 5.6 Describe two psychophysiological measures of Bourzac, 2016). However, there is conflicting evidence about brain activity. whether tES has beneficial effects on cognition; some stud- ies have even reported detrimental effects. Differing stimu- SCALP ELECTROENCEPHALOGRAPHY. The electroen- lation protocols might account for some of the discrepant cephalogram (EEG) is a measure of the gross electrical activ- findings (see Sellers et al., 2015). ity of the brain. It is recorded through large electrodes by a Transcranial ultrasound stimulation (tUS) is a tech- device called an electroencephalograph (EEG machine), and the nique that, like tES and TMS, can be used to activate partic- technique is called electroencephalography. In EEG studies ular brain structures. However, unlike tES and TMS, which of human participants, each channel of EEG activity is usu- can only be used to stimulate cortical structures, tUS can ally recorded from disk-shaped electrodes, about half the also be used to activate subcortical structures. size of a dime, which are attached to the scalp. To activate a brain structure using tUS, multiple sources The scalp EEG signal reflects the sum of electrical of low-amplitude ultrasonic sound waves are placed around events throughout the head. These events include action the head of the individual. Then, each of those sound potentials and postsynaptic potentials as well as electrical sources is directed at the target brain structure. When the signals from the skin, muscles, blood, and eyes. ultrasonic sound waves from each of those sources reach the Thus, the utility of the scalp EEG does not lie in its target structure they sum together, such that the amplitude ability to provide an unclouded view of neural activity. Its of the sound waves at the target brain structure is suffi- value as a research and diagnostic tool rests on the fact that ciently large to stimulate activity in the cells there (see Tyler, some EEG wave forms are associated with particular states Lani, & Hwang, 2018). of consciousness or particular types of cerebral pathology The tUS technique can also be used to make small (e.g., epilepsy). For example, alpha waves are regular, 8- to permanent lesions to a brain structure. The procedure is 12-per-second, high-amplitude waves that are associated the same as that for stimulation via tUS, except that the with relaxed wakefulness. A few examples of EEG wave amplitude of each ultrasound source is larger, leading to forms and their psychological correlates are presented in a larger amplitude waveform that is sufficient to create a Figure 5.8. small (e.g., the size of a grain of rice) permanent lesion. Because EEG signals decrease in amplitude as they This tUS-based lesion method has been used to treat sev- spread from their source, a comparison of signals recorded eral conditions (e.g., lesioning a thalamic nucleus to treat from various sites on the scalp can sometimes indicate essential tremor)—all without having to make an incision. the origin of particular waves (see Cohen, 2017). This is ­Accordingly, the tUS lesion technique is revolutionizing why it is usual to record EEG activity from many sites neurosurgery (see ­Landhuis, 2017). simultaneously. M05_PINE1933_11_GE_C05.indd 128 22/01/2021 10:52  The Research Methods of Biopsychology 129 A method used to reduce the noise of the background Figure 5.8 Some typical electroencephalograms and their psychological correlates. EEG is signal ­a veraging. First, a subject’s response to a stimulus, such as a click, is recorded many—let’s say 1,000—times. Then, a computer identifies the millivolt Aroused value of each of the 1,000 traces at its starting point (i.e., at the click) and calculates the mean of these 1,000 scores. Next, it considers the value of each of the 1,000 Relaxed Alpha waves traces 1 millisecond (msec) from its start, for example, and calculates the mean of these values. It repeats this process at the 2-msec mark, the 3-msec mark, and so on. When these averages are plotted, the average response Asleep evoked by the click is more apparent because the random background EEG is canceled out by the averaging. See Figure 5.9, which illustrates the averaging of an auditory evoked potential. Deep sleep The analysis of average evoked potentials (AEPs) focuses on the various waves in the averaged signal. Each wave is characterized by its direction, positive or negative, and by its latency. For example, the P300 wave illustrated in Figure 5.10 is the positive wave that occurs about 300 ­milliseconds after a momentary stimulus that has 1 second meaning for the participant (e.g., a stimulus to which the Psychophysiologists are often Figure 5.9 Signal averaging: Averaging of the background EEG (left) and of a­ uditory evoked potentials (right). Averaging increases the signal-to-noise ratio. more interested in the EEG waves that accompany certain psycholog- ical events than in the background 1-second segments 1-second segments EEG signal. These accompanying of background EEG of EEG with evoked EEG waves are generally referred potential to as e­vent-related potentials (ERPs). One commonly studied type of event-related potential is the sensory evoked potential— the change in the cortical EEG signal elicited by the momentary presentation of a sensory stimu- lus. As Figure 5.9 illustrates, the cortical EEG that follows a sensory stimulus has two components: the response to the stimulus (the sig- nal) and the ongoing background EEG activity (the noise). The signal is the part of any recording that is of interest; the noise is the part that isn’t. The problem in record- ing sensory evoked potentials is that the noise of the background EEG is often so great that the sen- sory evoked potential is masked. Average Average M easuring a sensory evoked ­ background evoked potential can be like detect- EEG potential Click ing a whisper at a rock concert. M05_PINE1933_11_GE_C05.indd 129 22/01/2021 10:52  130 Chapter 5 Figure 5.10 An average auditory evoked potential. Notice Figure 5.11 A magnetoencephalography (MEG) machine. the P300 wave. This wave occurs only if the stimulus has Stylish in any home! meaning for the participant; in this case, the ‘click’ sound signals the imminent delivery of a reward. By conven- tion, positive EEG waves are always shown as downward deflections. Far-field potentials P300 Meaningful click 200 400 600 Time (milliseconds) Image Source/Alamy Stock Photo participant must respond). In contrast, the portions of an motor neuron that innervates it. At any given time, a few evoked potential recorded in the first few milliseconds fibers in each resting muscle are likely to be contracting, after a stimulus are not influenced by the meaning of the thus maintaining the overall tone (tension) of the muscle. stimulus for the participant. These small waves are called Movement results when a large number of fibers contract far-field potentials because, although they are recorded at the same time. from the scalp, they originate far away in the sensory In everyday language, anxious people are commonly nuclei of the brain stem. referred to as “tense.” This usage acknowledges the fact that anxious or otherwise aroused individuals typically display MAGNETOENCEPHALOGRAPHY. Another technique high resting levels of tension in their muscles. This is why used to monitor brain activity from the scalp of human sub- psychophysiologists are interested in this measure; they use jects is magnetoencephalography (MEG). MEG measures it as an indicator of psychological arousal. changes in magnetic fields on the surface of the scalp that are Electromyography is the usual procedure for m ­ easuring produced by changes in underlying patterns of neural activ- muscle tension. The resulting record is called an electromyo- ity. Because the magnetic signals induced by neural activity gram (EMG). EMG activity is usually recorded between two are so small, only those induced near the surface of the brain electrodes taped to the surface of the skin over the muscle of can be recorded from the scalp (see Hari & ­Parkkonen, 2015). interest. An EMG record is presented in Figure 5.12. You will MEG has two major advantages over EEG. First, it notice from this figure that the main correlate of an increase has much better spatial resolution than EEG; that is, it can in muscle contraction is an increase in the amplitude of the localize changes in electrical activity in the cortex with raw EMG signal, which reflects the number of muscle fibers greater precision. Second, MEG can be used to localize sub- contracting at any one time. cortical activity with greater reliability than EEG (Baillet, Most psychophysiologists do not work with raw EMG 2017). Some downsides to the use of MEG include its high signals; they convert them to a more workable form. The price, the large size of the MEG machines (see Figure 5.11), raw signal is fed into a computer that calculates the total and the requirement that participants remain very still dur- amount of EMG spiking per unit of time—in consecutive ing recordings (Baillet, 2017; but see Boto et al., 2018). 0.1-second intervals, for example. The integrated signal (i.e., the total EMG activity per unit of time) is then plotted. Psychophysiological Measures of The result is a smooth curve, the amplitude of which is a Somatic Nervous System Activity simple, continuous measure of the level of muscle tension (see Figure 5.12). LO 5.7 Describe two psychophysiological measures of somatic nervous system activity. EYE MOVEMENT. The electrophysiological technique for recording eye movements is called ­electrooculography, MUSCLE TENSION. Each skeletal muscle is composed and the resulting record is called an electrooculogram (EOG). of millions of threadlike muscle fibers. Each muscle fiber ­E lectrooculography is based on the fact that a steady contracts in an all-or-none fashion when activated by the ­potential difference exists between the front (positive) and M05_PINE1933_11_GE_C05.indd 130 22/01/2021 10:52  The Research Methods of Biopsychology 131 Figure 5.12 The relation between a raw EMG signal Psychophysiological Measures of and its integrated version. The volunteer tensed her muscle Autonomic Nervous System Activity beneath the electrodes and then gradually relaxed it. LO 5.8 Describe two psychophysiological measures of autonomic nervous system activity. SKIN CONDUCTANCE. Emotional thoughts and experi- ences are associated with increases in the ability of the skin to conduct electricity. The two most commonly employed indexes of electrodermal activity are the skin ­conductance level (SCL) and the skin conductance response (SCR). The SCL is a measure of the background level of skin con- Raw EMG signal ductance that is associated with a particular situation, whereas the SCR is a measure of the transient changes in skin conductance that are associated with discrete experiences. The physiological bases of skin conductance changes are not fully understood, but there is considerable evidence implicating the sweat glands. Although the main function of sweat glands is to cool the body, these glands tend to become active in emotional situations, causing the release of sweat that in turn increases the electrical conductivity of Integrated EMG signal the skin (see Green et al., 2014). Sweat glands are distrib- uted over most of the body surface, but, as you are almost certainly aware, those of the hands, feet, armpits, and fore- back (negative) of the eyeball. Because of this steady poten- head are particularly responsive to emotional stimuli. tial, when the eye moves, a change in the electrical potential between electrodes placed around the eye can be recorded. CARDIOVASCULAR ACTIVITY. The presence in our It is usual to record EOG activity between two electrodes language of phrases such as white with fear and blushing placed on each side of the eye to measure its horizontal bride indicates that modern psychophysiologists were not movements and between two electrodes placed above the first to recognize the relationship between cardiovas- and below the eye to measure its vertical movements (see cular activity and emotion. The cardiovascular system has Figure 5.13). two parts: the blood vessels and the heart. It is a system for distributing oxygen and nutrients to the tissues of the body, removing metabolic wastes, and transmitting chemi- Figure 5.13 The typical placement of electrodes around cal messages. Three different measures of cardiovascular the eye for electrooculography. The two electrooculogram activity are frequently employed in psychophysiological traces were recorded as the volunteer scanned a circle. research: heart rate, arterial blood pressure, and local blood volume. Heart Rate The electrical signal associated with each heartbeat can be recorded through electrodes placed on the chest. The recording is called an electrocardiogram (abbre- viated either ECG, for obvious reasons, or EKG, from the original German). The average resting heart rate of a healthy adult is about 70 beats per minute, but it increases abruptly at the sound, or thought, of a dental drill. Blood Pressure Measuring arterial blood pressure involves two independent measurements: a measurement of the peak pressure during the periods of heart contraction, Electrooculograms of the participant as she scanned the systoles, and a measurement of the minimum pressure a circle. during the periods of relaxation, the diastoles. Blood pres- sure is usually expressed as a ratio of systolic over diastolic M05_PINE1933_11_GE_C05.indd 131 22/01/2021 10:52  132 Chapter 5 blood pressure in millimeters of mercury (mmHg). The experimental devices are precisely positioned in the depths normal resting blood pressure for an adult is about 130/70 of the brain. Two things are required in stereotaxic surgery: mmHg. A chronic blood pressure of more than 140/90 an atlas to provide directions to the target site and an instru- mmHg is viewed as a serious health hazard and is called ment for getting there. hypertension. The stereotaxic atlas is used to locate brain struc- You have likely had your blood pressure measured tures in much the same way that a geographic atlas is with a sphygmomanometer—a crude device composed of a used to locate geographic landmarks. There is, however, hollow cuff, a rubber bulb for inflating it, and a pressure one important difference. In contrast to the surface of the gauge for measuring the pressure in the cuff (sphygmos earth, which has only two dimensions, the brain has three. means “pulse”). More reliable, fully automated methods Accordingly, the brain is represented in a stereotaxic atlas are used in research. by a series of individual maps, one per page, each repre- senting the structure of a single, two-dimensional frontal Blood Volume Changes in the volume of blood in par- brain slice. In stereotaxic atlases, all distances are given in ticular parts of the body are associated with psychological millimeters from a designated reference point. In most rat events. The best-known example of such a change is the atlases, the reference point is bregma—the point on the top engorgement of the genitals associated with sexual arousal of the skull where two of the major sutures (seams in the in both males and females. Plethysmography refers to the skull) intersect. various techniques for measuring changes in the volume The stereotaxic instrument (see Figure 5.14) has two of blood in a particular part of the body (plethysmos means parts: a head holder, which firmly holds each subject’s brain “an enlargement”). in the prescribed position and orientation; and an electrode One method of measuring these changes is to record holder, which holds the device to be inserted. A system of the volume of the target tissue by wrapping a strain gauge precision gears allows the electrode holder to be moved in around it. Although this method has utility in measuring three dimensions: anterior–posterior, dorsal–ventral, and blood flow in fingers or similarly shaped organs, the pos- lateral–medial. The implantation by stereotaxic surgery sibilities for employing it are somewhat limited. Another of an electrode in the amygdala of a rat is illustrated in plethysmographic method is to shine a light through the Figure 5.15. tissue under investigation and to measure the amount of light absorbed by it. The more blood there is in a structure, the more light it will absorb. Figure 5.14 A stereotaxic instrument. This one is meant for surgery on rodents. Invasive Physiological Research Methods We turn now from a consideration of the noninvasive techniques employed in research on living human brains to a consideration of more direct techniques, which are commonly employed in biopsychological studies of non- human animals. Most physiological techniques used in biopsychological research on nonhuman animals fall into one of three categories: lesion methods, electrical stimu- lation methods, and invasive recording methods. Each of these three methods is discussed in this module, but we begin with a description of stereotaxic surgery because each of these methods involves the use of stereotaxic surgery. Stereotaxic Surgery LO 5.9 Describe the process of stereotaxic surgery. Stereotaxic surgery is the first step in many biopsychologi- Model 900 Small Animal Stereotaxic Instrument originally designed by David cal experiments. Stereotaxic surgery is the means by which Kopf Instruments in 1963. M05_PINE1933_11_GE_C05.indd 132 22/01/2021 10:52  The Research Methods of Biopsychology 133 cortical tissue itself, a skilled surgeon can Figure 5.15 Stereotaxic surgery: Implanting an electrode in the rat amygdala. delicately peel off the layers of cortical tis- sue from the surface of the brain, leaving 0 7 6 5 4 3 2 1 0 1 2 3 4 5 6 7 Bregma 2.8 0 1 The stereotaxic atlas indicates that the amygdala target site is 5 mm ventral. This page of the atlas the underlying white matter and major 1 2 1 2 blood vessels undamaged. 3 3 represents a frontal section that is 4 4 2.8 mm posterior to bregma. RADIO-FREQUENCY LESIONS. Small 5 5 6 7 6 7 subcortical lesions are commonly made 8 9 8 9 by passing radio-frequency current (high- 10 10 frequency current) through the target 7 6 5 4 3 2 1 0 1 2 3 4 5 6 7 tissue from the tip of a stereotaxically Amygdala positioned electrode. The heat from the Bregma current destroys the tissue. The size and 2 A hole is drilled 2.8 mm posterior to bregma and 4.5 mm lateral to it. Then, the electrode shape of the lesion are determined by the duration and intensity of the current and holder is positioned over the hole, the configuration of the electrode tip. and the electrode is lowered 8.5 mm through the hole (i.e., 8.5 mm KNIFE CUTS. Sectioning (cutting) is used ventral). to eliminate conduction in a nerve or tract. A tiny, well-placed cut can unambiguously accomplish this task without producing Dental acrylic extensive damage to surrounding tissue. Electrode connector Electrode 3 The electrode is anchored to the skull with several stainless steel screws and dental acrylic that How does one insert a knife into the brain to make a cut without severely damag- is allowed to harden around the ing the overlying tissue? One method is electrode connector. depicted in Figure 5.16. REVERSIBLE LESIONS. Reversible lesions are useful alternatives to destruc- tive lesions. Reversible lesions are methods for temporarily eliminating the activity in a particular area of the brain while tests are being conducted. The advantage of reversible lesions is that the same sub- jects can be repeatedly tested in both Lesion Methods the lesion and control conditions. The two most common methods of producing a reversible lesion are by temporar- LO 5.10 Describe four types of lesion methods and ily cooling the target structure or by injecting an anesthetic explain why it is important to be cautious when (e.g., lidocaine) into it. interpreting the effects of lesions. INTERPRETING LESION EFFECTS. Before you leave this Those of you with an unrelenting drive to dismantle objects section on lesions, a word of caution is in order. Lesion to see how they work will appreciate the lesion methods. In effects are deceptively difficult to interpret. Because the those methods, a part of the brain is damaged, destroyed, structures of the brain are small, convoluted, and tightly or inactivated; then the behavior of the subject is care- packed together, even a highly skilled surgeon cannot com- fully assessed in an effort to determine the functions of the pletely destroy a structure without producing significant lesioned structure. Four types of lesions are discussed here: damage to adjacent structures. There is, however, an unfor- aspiration lesions, radio-frequency lesions, knife cuts, and tunate tendency to lose sight of this fact. For example, a reversible lesions. lesion that leaves major portions of the amygdala intact and ASPIRATION LESIONS. When a lesion is to be made in an damages an assortment of neighboring structures comes to area of cortical tissue that is accessible to the eyes and instru- be thought of simplistically as an amygdala lesion. Such an ments of the surgeon, aspiration is frequently the method of apparently harmless abstraction can be misleading in two choice. The cortical tissue is drawn off by suction through a ways. If you believe that all lesions referred to as “amyg- fine-tipped handheld glass pipette. Because the underlying dala lesions” include damage to no other brain structure, white matter is slightly more resistant to suction than the you may incorrectly attribute all of their behavioral effects M05_PINE1933_11_GE_C05.indd 133 22/01/2021 10:52  134 Chapter 5 immediate increase in the firing of neurons near the tip of Figure 5.16 A device for performing subcortical knife cuts. The device is stereotaxically positioned in the brain; the electrode. then the blade swings out to make the cut. Here, the anterior Electrical stimulation of the brain is an important biopsy- commissure is being sectioned. chological research tool because it often has behavioral effects, usually opposite to those produced by a lesion to the same site. It can elicit a number of behavioral sequences, including eating, drinking, attacking, copulating, and sleeping. The par- ticular behavioral response elicited depends on the location of the electrode tip, the parameters of the current, and the test environment in which the stimulation is administered. Because electrical stimulation of the brain is an inva- sive procedure, its use is usually limited to nonhumans. ­However, sometimes the brains of conscious human patients are stimulated for therapeutic reasons (e.g., Jonas et al., 2014). Invasive Electrophysiological Recording Methods LO 5.12 Describe four invasive electrophysiological recording methods. This section describes four invasive electrophysiological recording methods: intracellular unit recording, extracellular unit recording, multiple-unit recording, and invasive EEG recording. See Figure 5.17 for an example of each method. INTRACELLULAR UNIT RECORDING. This method provides a moment-by-moment record of the graded fluc- tuations in one neuron’s membrane potential. Most experi- to amygdala damage; conversely, if you believe that all ments using this recording procedure are performed on lesions referred to as “amygdala lesions” include the entire chemically immobilized animals because it is difficult to amygdala, you may incorrectly conclude that the amyg- keep the tip of a microelectrode positioned inside a neuron dala does not participate in behaviors uninfluenced by the of a freely moving animal (see Long & Lee, 2012). However, lesion. special electrodes are now being developed that can allow BILATERAL AND UNILATERAL LESIONS. As a general researchers to do intracellular recordings in a freely moving principle—but one with several notable exceptions—the animal (see Lee & Brecht, 2018). behavioral effects of unilateral lesions (lesions restricted to EXTRACELLULAR UNIT RECORDING. With extracel- one half of the brain) are much milder than those of sym- lular unit recording, it is possible to record the activity metrical bilateral lesions (lesions involving both sides of the of a neuron through a microelectrode whose tip is posi- brain), particularly in nonhuman species. Indeed, behav- tioned in the extracellular fluid next to it—each time the ioral effects of unilateral lesions to some brain structures neuron fires, there is an electrical disturbance and a blip can be difficult to detect. As a result, most experimental is recorded at the electrode tip. Accordingly, extracellular studies of lesion effects are studies of bilateral, rather than unit recording provides a record of the firing of a neu- unilateral, lesions. ron but no information about the neuron’s membrane potential. It is difficult to record extracellularly from a Electrical Stimulation single neuron in a freely moving animal without the elec- trode tip shifting away from that neuron, but it can be LO 5.11 Describe the technique of electrical brain accomplished with special flexible microelectrodes that stimulation. can shift slightly with the brain. Initially, extracellular Clues about the function of a neural structure can be unit recording involved recording from one neuron at a obtained by stimulating it electrically. Electrical brain stim- time, each at the tip of a separately implanted electrode. ulation is usually delivered across the two tips of a bipo- However, it is now possible to simultaneously record lar electrode—two insulated wires wound tightly together extracellular signals from up to about 1,000 neurons by and cut at the end. Weak pulses of current produce an analyzing the correlations among the signals picked up M05_PINE1933_11_GE_C05.indd 134 22/01/2021 10:52  The Research Methods of Biopsychology 135 Figure 5.17 Four methods of recording electrical activity of the nervous system. An Intracellular Unit Recording A Multiple-Unit Recording An intracellular microelectrode records the A small electrode records the action potentials of membrane potential from one neuron as it fires. many nearby neurons. These are added up and plotted. In this example, firing in the area of the electrode tip gradually declined and then suddenly increased. Action Potentials (per millisecond) Membrane Number of (millivolts) Potential 1 2 3 4 5 10 20 30 40 50 Milliseconds Milliseconds Seconds An Extracellular Unit Recording An Invasive EEG Recording An extracellular microelectrode records the electrical A large implanted electrode picks up general disturbance that is created each time an adjacent neuron changes in electrical brain activity. The EEG signal fires. In this example, each vertical line represents an is not related to neural firing in any obvious way. action potential. (microvolts) (millivolts) Voltage Voltage 10 20 30 40 50 1 2 3 4 5 Milliseconds Seconds through several different electrodes implanted in the same general area (see Callaway & Garg, 2017; Harris et al., 2016; Jun et al., 2017). Pharmacological Research MULTIPLE-UNIT RECORDING. In multiple-unit record- Methods ing, the electrode tip is much larger than that of a micro- In the preceding module, you learned how physiologi- electrode; thus, it picks up signals from many neurons, and cal psychologists study the brain by manipulating it and slight shifts in its position due to movement of the subject recording from it using surgical and electrical methods. have little effect on the overall signal. The many action In this module, you will learn how psychopharmacolo- potentials picked up by the electrode are fed into an inte- gists manipulate the brain and record from it using chem- grating circuit, which adds them together. A multiple-unit ical methods. recording is a graph of the total number of recorded action The major research strategy of psychopharmacology potentials per unit of time (e.g., per 0.1 second). is to administer drugs that either increase or decrease the INVASIVE EEG RECORDING. In nonhuman animals, and effects of particular neurotransmitters and to observe the sometimes in human patients (see Fox et al., 2018), EEG sig- behavioral consequences. Described here are routes of nals can be recorded through implanted electrodes rather drug administration, methods of using chemicals to make than through scalp electrodes. In nonhuman animals, corti- selective brain lesions, methods of measuring the chemical cal EEG signals are frequently recorded through stainless activity of the brain that are particularly useful in biopsy- steel skull screws, whereas subcortical EEG signals are typi- chological research, and methods for locating neurotrans- cally recorded through implanted wire electrodes. mitter systems. M05_PINE1933_11_GE_C05.indd 135 22/01/2021 10:52  136 Chapter 5 Routes of Drug Administration proved particularly useful in biopsychological research are the 2-deoxyglucose technique and cerebral dialysis. LO 5.13 Describe the various methods of drug 2-DEOXYGLUCOSE TECHNIQUE. The 2-deoxyglucose administration. (2-DG) technique entails placing an animal that has been In most psychopharmacological experiments, drugs are injected with radioactive 2-DG in a test situation in which administered in one of the following ways: (1) they are fed it engages in an activity of interest. Because 2-DG is simi- to the subject; (2) they are injected through a tube into the lar in structure to glucose—the brain’s main source of stomach (intragastrically); or (3) they are injected hypoder- energy—neurons active during the test absorb it at a high mically into the peritoneal cavity of the abdomen (intraperi- rate but do not metabolize it. Then the subject is killed, and toneally, IP), into a large muscle (intramuscularly, IM), into its brain is removed and sliced. The slices are then sub- the fatty tissue beneath the skin (subcutaneously, SC), or into jected to ­autoradiography: They are coated with a photo- a large surface vein (intravenously, IV). A problem with these graphic emulsion, stored in the dark for a few days, and peripheral routes of administration is that many drugs do then developed much like film. Areas of the brain that not readily pass through the blood–brain barrier. To over- absorbed high levels of radioactive 2-DG during the test come this problem, drugs can be administered in small appear as black spots on the slides. The density of the spots amounts through a fine, hollow tube, called a c­ annula, that in various regions of the brain can then be color-coded (see has been stereotaxically implanted in the brain. Figure 5.18). CEREBRAL DIALYSIS. Cerebral dialysis is a method of Selective Chemical Lesions measuring the extracellular concentration of specific neu- LO 5.14 Describe the method of selective neurotoxic rochemicals in behaving animals (most other techniques lesions. for measuring neurochemicals require that the subjects be killed so that tissue can be extracted). Cerebral dialysis The effects of surgical, radio-frequency, and reversible involves implanting a fine tube with a short semipermeable lesions are frequently difficult to interpret because they affect all neurons in the target area. In some cases, it is possible to make Figure 5.18 The 2-deoxyglucose technique. The accumulation of more selective lesions by injecting neuro- ­radioactivity is shown in three frontal sections taken from the brain of a toxins (neural poisons) that have an affinity ­Richardson’s ground squirrel. The subject was injected with radioactive 2-deoxyglucose; then, for 45 minutes, it viewed brightly illuminated black and for certain components of the nervous sys- white stripes through its left eye while its right eye was covered. Because tem. There are many selective neurotoxins. the ground squirrel visual system is largely crossed, most of the radioactivity For example, when either kainic acid or ibo- accumulated in the visual structures of the right hemisphere. tenic acid is administered by microinjection, Left Right it is preferentially taken up by cell bodies at the tip of the cannula and destroys those neurons, while leaving neurons with axons passing through the area largely unscathed. Another selective neurotoxin that has been widely used is 6-hydroxydopamine (6-OHDA). It is taken up by only those neu- rons that release the neurotransmitter nor- epinephrine or dopamine, and it leaves other neurons at the injection site undamaged. Measuring Chemical Activity of the Brain LO 5.15 Describe two techniques for measuring chemical activity in the brain. There are many procedures for measuring the chemical activity of the brains of labo- ratory animals. Two techniques that have Rod Cooper/University of Calgary Department of Psychology M05_PINE1933_11_GE_C05.indd 136 22/01/2021 10:52  The Research Methods of Biopsychology 137 section into the brain. The semi- Figure 5.19 Immunocytochemistry. This section through a rat’s pons reveals nor- permeable section is positioned in adrenergic neurons that have attracted the antibody for dopamine-beta-hydroxylase, the brain structure of interest so the enzyme that converts dopamine to norepinephrine. that extracellular chemicals from the structure will diffuse into the tube. Once in the tube, they can be collected for freezing, storage, and later analysis; or they can be carried in solution directly to a chromatograph (a device for mea- suring the chemical constituents of liquids or gases). Locating Neurotransmitters and Receptors in the Brain LO 5.16 Describe two techniques for locating particular neurotransmitters or receptors in the brain. A key step in trying to understand Richard Mooney, University of Toledo College of Medicine, Department of Neurosciences the psychological function of a par- ticular neurotransmitt