Name: preterm eeg: a multimodal neurophysiological protocol
Uploaded: 2012-02-18T14:00:00
Duration: 19 min 32 s
Description: Watch this Scientific Journal Video about Preterm EEG: A Multimodal Neurophysiological Protocol at JoVE.com

We want to thank Mr. Jyri Ojala for the technical production of the film, including its animations, graphical design, as well as all technical editing. We also want to thank the parents who gave permission to have their baby be starring in this video, as well as the nurses (especially Mr. Jarmo M&auml;ki) for the help in preparation of film production. This work has received practical and/or financial support from Helsinki University Hospital, Helsinki University of Applied Sciences (Metropolia), Juselius Foundation, Erkko Foundation, as well as the Foundation of Pediatrics (Lastentautien tutkimuss&auml;&auml;ti&ouml;) and from the European Community's Seventh Framework Programme European Community FP7-PEOPLE-2009-IOF, grant agreement n&deg;254235.
The manufacturer of the dense array EEG caps suitable for recording preterm babies is ANT B.V. For contact information, please visit <a href="http://www.ant-neuro.com">http://www.ant-neuro.com</a>. Notably, the NicOne EEG amplifier (CareFusion, Madison, USA) used in our video is not a genuine DC-coupled amplifier. Following manufacturers do currently (October, 2011) provide DC-coupled, clinically suitable EEG amplifiers: CareFusion (their newest amplifier), ANT Neuro (<a href="http://www.ant-neuro.com">www.ant-neuro.com</a>), EGI (<a href="http://www.egi.com">www.egi.com</a>), BrainProducts (<a href="http://www.BrainProducts.com">www.BrainProducts.com</a>), Neuroscan (<a href="http://www.neuroscan.com">www.neuroscan.com</a>), as well as SACS (<a href="http://www.sacs.se">www.sacs.se</a>).

1. Preparing the baby and the cot/incubator for an EEG study
<ol>
 <li>Schedule the EEG study so that it does not co-incide with the other NICU procedures, such as blood sampling or ultrasound examination, as they may interfere with both the EEG devices and baby's vigilance state. </li>
 <li>Start recording immediately after feeding, so the baby is most likely to fall into sleep during EEG recording. It may be practical to apply all recording electrodes prior to feeding and/or other care procedures, so that the EEG can commence immediately thereafter.</li>
 <li>Make sure that there are no unnecessary electric devices connected to the baby.</li>
 <li>Place all wires related to the EEG recording as far apart as possible from the other wires in the cot. Turn off bed heating, if possible treatment wise.</li>
 <li>In a case of electrical interference, you may try to identify the interfering device by turning them off (sometimes even unplugging from mains one at a time may be helpful).</li>
 <li>Make sure that baby's head is not touching damp or wet textiles, which may increase coupling of interference from external noise sources to the baby.</li>
</ol>
2. Application of the dense array EEG cap
<ol>
 <li>Remove interfering wax and oil from baby's scalp by swiping it with a cloth wetted with diluted alcohol or baby shampoo. Assess visually the head size of the baby to select the optimal size of the EEG cap in order to make sure that the cap sits firmly at vertex, and that the temporal electrodes are located at an appropriate level above ear lobes (five different sizes of these caps are available to fit from extremely young preterm to fullterm neonates).</li>
 <li>If possible, two persons should work together to place the EEG cap, one taking care of the cap, and the other one holding baby's head and possible intubation or nasal CPAP tubes. If the patient has a CPAP, remove it for the 10-20 seconds that it takes to place the EEG cap. The holding straps of the nasal CPAP are wrapped over the EEG cap later.</li>
 <li>(optional) If the baby has an iv line in the head, you will need to disconnect it, and pass it through the outlet made for it in the cap (available in the caps used in the present study). Otherwise, you will need to use head net and single electrode placement, which is demonstrated on the website of the NEMO study (see <a href="http://www.nemo-europe.com/en/educational-tools.php">http://www.nemo-europe.com/en/educational-tools.php</a>).</li>
 <li>Hold the baby so that his/her face is directed towards you. Take the cap into both hands, and by using your fingers, push the middle of the cap inside out, so that the central (Cz and Pz electrodes) will first touch the scalp.</li>
 <li>Roll the cap gently over the head so that the cap is almost sitting in its final position. Finally, adjust the cap symmetrically in both left-right and anterior-posterior directions. Check symmetry by making sure that the Cz electrode is in the middle (left-right) at the line that connects both ears. When pulling the cap over forehead, it is practical to keep baby's head in place by pressing your thumbs against his/her forehead above the eyes.</li>
 <li>Fasten the chin strap to a tightness that keeps the cap well in place, and yet allows unrestricted breathing. Do also check now that the cap size is optimal. The cap shown in this video may look little loose. However, it extends well over the forehead and occipital areas, and yet it is well enough in contact with the scalp at centroparietal areas. Five different sizes of these caps are available to fit from extremely young preterm to full-term neonates, and the cap shown in this video will fit this baby for a few weeks from now on.</li>
 <li>Apply the electrode gel through the electrode holes with a syringe connected to a thick blunt gel needle ("tip"). A bended tip may be useful to reach contacts that are difficult to access directly. Fill the electrode cups so that no gel will leak out, as it may form bridges with neighbouring electrodes or even wet the pillow possibly causing artefact. A more viscous gel formula may be practical in this regard, and allows even gelling before cap placement (see ref 1).</li>
 <li>Check the impedances and EEG signal quality, and prepare the scalp either by gentle mechanical rubbing or by using the so called SurePrep method as needed2,3 (<a href="http://www.helsinki.fi/science/eeg/videos/sureprep/">http://www.helsinki.fi/science/eeg/videos/sureprep/</a>). Observe the quality of skin contact simultaneously from the impedance values given by the EEG software, or by looking at the EEG trace quality. Start from the reference and ground, then prepare one hemisphere at a time to minimize moving of the baby.</li>
 <li>Start EEG recording when the impedance levels are acceptable, usually below 15kohms, or when the signal does look clean enough without filtering. Note that impedances often improve over time, so give some minutes time for the impedances to settle down.</li>
</ol>
3. Application of the polygraphic sensors 
<ol>
 <li>Record muscle tone (electromyography; EMG) by attaching two EMG electrodes under the chin. Prepare the skin gently before electrode application.</li>
 <li>Record heart beat by applying two ECG electrodes over the chest or on the shoulders. Shoulder attachment is useful if there is a need to identify potential epileptic body movements. Prepare the skin gently before electrode application.</li>
 <li>Record eye movements (electro-oculogram, EOG) either with frontal EEG electrodes (then no additional electrodes are needed), or with separate EOG electrodes attached near the lateral corners of the eyes. In the youngest premies, however, it may be sometimes needed to use piezo sensors attached to eyelids (not shown in this video), because their retina may not be polarized enough to cause detectable electric fields. Prepare the skin gently before electrode application.</li>
 <li>Monitor respiration movements with a stretch-sensitive transducer belt that is wrapped around the trunk over abdomen or chest. Make sure that the transducer is tight enough to show respiratory movements in the recording, and yet loose enough to allow unrestricted respiration. In babies with spontaneous breathing, special care is needed not to restrain breathing movements (see also <a href="http://www.nemo-europe.com/en/educational-tools.php">http://www.nemo-europe.com/en/educational-tools.php</a>).</li>
 <li>If clinically needed, add also a movement-sensitive piezo sensor to any body part that is expected to show unusual movements.</li>
 <li>Position the EEG-synchronized video camera so that the whole baby is well seen in the picture. During specific parts of the EEG study, one may choose to focus on selected body parts, such as hands or feet during their tactile stimulation.</li>
</ol>
4. Recording 
<ol>
 <li>During the entire recording session, keep an eye on the signal quality, and make corrections if any signal deteriorates. Follow the baby as well, and add annotations of any event (eg. clinical sign, movement, drug administration) that may be of interest during later offline EEG interpretation.</li>
 <li>Should artefacts occur during the recording, troubleshoot them and make corrections to the recording immediately. Many artefacts may permanently compromise the later use of EEG signal.</li>
 <li>Adjust the recording length individually so that the EEG study includes at least one epoch of all vigilance states: awake, active sleep and quiet sleep. In most cases, this will take 40 to 90 minutes.</li>
 <li>Perform sensory stimulations (see section 5 below) when the baby is well asleep. It is preferable to wait until quiet sleep period. </li>
 <li>In the end of the EEG recording session, remove the cap and other sensors, and clean the excess gel from baby's scalp by wiping with a wet cloth.</li>
 <li>Clean the EEG cap and other accessories first mechanically, then sterilize them according to hospital instructions.</li>
</ol>
5. Sensory stimulations 
<ol>
 <li>Note that all sensory responses in the young preemies have a long (several seconds) refractory period, and the responses decline rapidly with short interstimulus intervals or in the presence of other ongoing EEG activity in the sensory cortex. Hence, deliver sensory stimuli at moments when the EEG has been relatively silent for at least few seconds, which is easiest to do during quiet sleep exhibiting trace discontinue activity4-6. If possible, use stimulation devices that automatically generate a mark (trigger) into the EEG trace. Perform first the tactile stimulations, then the visual stimuli, and at last the auditory stimuli, which may wake up the baby.</li>
 <li>To study the visual evoked responses, give single flashes with a device integrated to the EEG system. You may also show any transient light, such as moving a torch beam over the face. A response is readily generated even from distance, such as through the transparent incubator walls. Observe the response with at least one second duration in several occipital and parietal electrodes.</li>
 <li>To study the somatosensory evoked responses, apply gentle tactile stimuli to the palm or sole of the baby. It is helpful to use a device that automatically generates a mark (trigger) into the EEG trace. Observe the response in the contralateral centro-temporal (C and T) electrodes after hand stimulation, and in the midline (Cz) electrodes after foot stimulation. To assist later analysis, add manual annotation to EEG recording any time when you see a spontaneous limb movement, because it produces responses similar to stimulated ones. </li>
 <li>To study auditory responses, you may use almost any sound with relatively low intensity,such as hand clapping near baby's head. Avoid using the traditional horn stimulation, because its high sound intensity will lead to both auditory activation and sleep arousal, with often ensuing movement artefacts.</li>
</ol>
6. Analysis
<ol>
 <li>Start preliminary analysis during the EEG recording already. Add annotations of clinical events that are not obvious from the EEG signal (eg. care procedures, unusual movements or drug administration). Make note also of all EEG artefacts (eg. sucking, rocking or tapping), which are difficult to identify afterwards. Finally, add your preliminary considerations as they arise online, as they may often involve insights that are difficult to perceive later. Make sure that you have recorded both active and quiet sleep, and that the number and quality of sensory stimuli are sufficient.</li>
 <li>The actual, thorough review of neonatal EEG is done offline in a workstation where EEG findings can be processed and described in more detail. </li>
</ol>
7. Representative Results
<img src="/files/ftp_upload/3774/3774fig1.jpg" alt="Figure 1" /> 
Figure 1. Comparison of the recording with conventional EEG (left) and an unfiltered, Full-band EEG (FbEEG; right). Note the prominent slow fluctuations in the FbEEG signal [see also refs 7-9], which are absent in the conventional EEG. For details, see Discussion. 
<img src="/files/ftp_upload/3774/3774fig2.jpg" alt="Figure 2" /> 
Figure 2. Comparison of the increase in information obtained by adding the number of electrodes. A high density EEG (right) makes it possible to analyse and/or follow cortical areas separately. This spatial information is negligible in the conventional 8 channel (middle) recording, and completely lost in the common one channel EEG monitoring (left). Please <a href="https://www.jove.com/files/ftp_upload/3774/3774fig2_lg.jpg">click here</a> to see a larger version of this figure.
<img src="/files/ftp_upload/3774/3774fig3.jpg" alt="Figure 3" /> 
Figure 3. Left: An example is shown from single trial responses to tactile sensory stimulations of hands as seen in the raw EEG trace. Right: Comparison of the preterm and fullterm somatosensory responses (both C4-Fz derivation) demonstrates the magnitude of preterm cortical reactions. The preterm response is shown from a single trial trace, while the fullterm response is generated by averaging, since it would not be distinguished at single trial level. In the fullterm trace, the arrow depicts the N1 response that is conventionally used as the representative measure for clinical diagnosis. For further details, see refs 4,6,10,11. Please <a href="https://www.jove.com/files/ftp_upload/3774/3774fig3_lg.jpg">click here</a> to see a larger version of this figure.

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No conflicts of interest declared.

Recording of neonatal EEG in the way shown here is safe, and hence doable from any baby and in any condition that allows handling associated with routine care procedures1. The intensive care unit is a challenging environment to the sensitive EEG devices. A key to a technically good quality recording is a proper use of well-functioning dense array EEG cap, such as the one shown in this video. NICU environment is unique in that the study subjects are critically ill, vulnerable babies, which undergo demanding care and diagnostic procedures. To ensure not only safety of the patient, but also the future of EEG studies in your NICU, it is necessary to have a close collaboration with your key NICU contact, as well as a high level of trust with all NICU staff involved in patient care. 
Recent work in both human babies5-7,10 and animal models with rat pups13-15 has emphasized the dominance of infraslow frequencies in the EEG. While they are readily seen with the FbEEG technique (see Fig 1 and refs 8,9), they are ignored or distorted in the conventional EEG (using an AC coupled amplifier) that permanently cuts them at the time of signal collection, because AC coupled amplifiers act as highpass (low cut) filters. A faithful recording of these infraslow activities requires a DC stable recording setting that consists of a DC coupled amplifier (see Acknowledgements), Ag/AgCl electrodes, chloride-containing gel16, as well as an adequate elimination of epithelial potential (for details, see refs 2,3,8, as well as <a href="http://www.nemo-europe.com/en/educational-tools.php">http://www.nemo-europe.com/en/educational-tools.php</a> -&gt; Setting up EEG hardware -&gt; Skin prepping). It is notable in clinical context, that the caps used in our presentation are fully compliant with FbEEG recording, while many other clinical EEG caps are unsuitable because of their inappropriate electrode materials (eg. tin) 16 and/or poor mechanical stability due to the cut and electrode holder design (for details, see also refs 1,8).
Adequate assessment or monitoring of the early preterm brain must be based on a thorough understanding of the nature and specific characteristics of the immature brain activity itself (see above). Such an approach is largely lacking in the current clinical practice and literature. To meet this need, the protocol for a multimodal neurophysiological assessment of preterm babies was developed in our laboratory. The present paper bridges together the latest knowledge of developmental neurobiology, the relevant advances in neurophysiological techniques, as well as the pressing need for novel clinical research. Our work hences opens the window for translational studies to be performed in a bidirectional manner (from bench to bedside and back). In addition, clear demonstration of the protocol aims to open a venue for larger scale collection of clinically relevant datasets, including studies to define the elusive criteria of normality.
Our clinical experience in the Helsinki University Hospital has shown that i) multimodal studies of this kind have rapidly become an integral part of
clinical routine, ii) that they have significantly increased the interest in neonatal EEG studies, and iii) that the developments in recording
techniques shown here have made performance of such recordings as readily attainable as any limited, conventional EEG. Most importantly, understanding the relationships between developing brain function and structure makes it possible to assess brain maturation at a time when the baby is not yet able to communicate with the outside world 17. Better brain care at an early stage of development is likely to lead to a permanent increase in health and in overall quality of life of the preterm baby.

Links for the SurePrep method
<a href="http://www.helsinki.fi/science/eeg/videos/sureprep/SurePrep_for_SP_paper.mov">http://www.helsinki.fi/science/eeg/videos/sureprep/SurePrep_for_SP_paper.mov</a>
<a href="http://www.helsinki.fi/science/eeg/videos/sureprep/SurePrep_for_NEMO.wmv">http://www.helsinki.fi/science/eeg/videos/sureprep/SurePrep_for_NEMO.wmv</a>
Online video link to additional demonstrations on neonatal EEG recording
<a href="http://www.nemo-europe.com/en/educational-tools-for-clinicians-and-health-care-professionals.php">http://www.nemo-europe.com/en/educational-tools-for-clinicians-and-health-care-professionals.php</a>
 Online video link to the theory and practice of scalp preparation
 <a href="http://www.helsinki.fi/science/eeg/videos/sureprep/Nemo_hardware_Scalp_prepping.f4v">http://www.helsinki.fi/science/eeg/videos/sureprep/Nemo_hardware_Scalp_prepping.f4v</a>

preterm eeg: a multimodal neurophysiological protocol

Since its introduction in early 1950s, electroencephalography (EEG) has been widely used in the neonatal intensive care units (NICU) for assessment and monitoring of brain function in preterm and term babies. Most common indications are the diagnosis of epileptic seizures, assessment of brain maturity, and recovery from hypoxic-ischemic events. EEG recording techniques and the understanding of neonatal EEG signals have dramatically improved, but these advances have been slow to penetrate through the clinical traditions. The aim of this presentation is to bring theory and practice of advanced EEG recording available for neonatal units. 
In the theoretical part, we will present animations to illustrate how a preterm brain gives rise to spontaneous and evoked EEG activities, both of which are unique to this developmental phase, as well as crucial for a proper brain maturation. Recent animal work has shown that the structural brain development is clearly reflected in early EEG activity. Most important structures in this regard are the growing long range connections and the transient cortical structure, subplate. Sensory stimuli in a preterm baby will generate responses that are seen at a single trial level, and they have underpinnings in the subplate-cortex interaction. This brings neonatal EEG readily into a multimodal study, where EEG is not only recording cortical function, but it also tests subplate function via different sensory modalities. Finally, introduction of clinically suitable dense array EEG caps, as well as amplifiers capable of recording low frequencies, have disclosed multitude of brain activities that have as yet been overlooked.
In the practical part of this video, we show how a multimodal, dense array EEG study is performed in neonatal intensive care unit from a preterm baby in the incubator. The video demonstrates preparation of the baby and incubator, application of the EEG cap, and performance of the sensory stimulations.

Watch this Scientific Journal Video about Preterm EEG: A Multimodal Neurophysiological Protocol at JoVE.com

Preterm EEG: A Multimodal Neurophysiological Protocol

This video explains the background theory of the neonatal EEG activity and the sensory responses, followed by a live demonstration of their recording in neonatal intensive care unit.

Since its introduction in early 1950s, electroencephalography (EEG) has been widely used in the neonatal intensive care units (NICU) for assessment and ...

preterm-eeg-a-multimodal-neurophysiological-protocol

Research

JoVE Journal

Neuroscience

Functional Mapping with Simultaneous MEG and EEG

We use magnetoencephalography (MEG) and electroencephalography (EEG) to locate and determine the temporal evolution in brain areas involved in the processing of simple sensory stimuli. We will use somatosensory stimuli to locate the hand somatosensory areas, auditory stimuli to locate the auditory cortices, visual stimuli in four quadrants of the visual field to locate the early visual areas. These type of experiments are used for functional mapping in epileptic and brain tumor patients to locate eloquent cortices. In basic neuroscience similar experimental protocols are used to study the orchestration of cortical activity. The acquisition protocol includes quality assurance procedures, subject preparation for the combined MEG/EEG study, and acquisition of evoked-response data with somatosensory, auditory, and visual stimuli. We also demonstrate analysis of the data using the equivalent current dipole model and cortically-constrained minimum-norm estimates. Anatomical MRI data are employed in the analysis for visualization and for deriving boundaries of tissue boundaries for forward modeling and cortical location and orientation constraints for the minimum-norm estimates.

We use magnetoencephalography (MEG) and electroencephalography (EEG) to map brain areas involved in the processing of simple sensory stimuli.

We use magnetoencephalography (MEG) and electroencephalography (EEG) to locate and determine the temporal evolution in brain areas involved in the ...

Non-restraining EEG Radiotelemetry: Epidural and Deep Intracerebral Stereotaxic EEG Electrode Placement

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and neurodegenerative diseases as well as epilepsies. This powerful technique does not only provide valuable insights into the underlying pathophysiological mechanisms, i.e., the etiopathogenesis of CNS related diseases, it also facilitates the development of new translational, i.e., therapeutic approaches. Whereas competing techniques that make use of recorder systems used in jackets or tethered systems suffer from their unphysiological restraining to semi-restraining character, radiotelemetric EEG recordings overcome these disadvantages. Technically, implantable EEG radiotelemetry allows for precise and highly sensitive measurement of epidural and deep, intracerebral EEGs under various physiological and pathophysiological conditions. First, we present a detailed protocol of a straight forward, successful, quick and efficient technique for epidural (surface) EEG recordings resulting in high-quality electrocorticograms. Second, we demonstrate how to implant deep, intracerebral EEG electrodes, e.g., in the hippocampus (electrohippocampogram). For both approaches, a computerized 3D stereotaxic electrode implantation system is used. The radiofrequency transmitter itself is implanted into a subcutaneous pouch in both mice and rats. Special attention also has to be paid to pre-, peri- and postoperative treatment of the experimental animals. Preoperative preparation of mice and rats, suitable anesthesia as well as postoperative treatment and pain management are described in detail.

Non-restraining EEG radiotelemetry is a valuable methodological approach to record in vivo long-term electroencephalograms from freely moving rodents. This detailed protocol describes stereotaxic epidural and deep intracerebral electrode placement in different brain regions in order to obtain reliable recordings of CNS rhythmicity and CNS related behavioral stages.

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and ...

Recording EEG in Freely Moving Neonatal Rats Using a Novel Method

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such as epilepsy and neurodegenerative disorders. However, it is technically difficult to obtain EEG recordings in neonates as it requires specialized handling and great care. Here, we present a novel method to record EEG in neonatal rat pups (P8-P15). We designed a simple and reliable electrode using computer pin loci; it can be easily implanted into the skull of a rat pup to record high-quality EEG signals in the normal and epileptic brain. Pups were given an intraperitoneal (i.p.) injection of the neurotoxin kainic acid (KA) to induce epileptic seizures. The surgical implantation performed in this procedure is less expensive than other EEG procedures for neonates. This method allows one to record high-quality and stable EEG signals for more than 1 week. Furthermore, this procedure can also be applied to adult rats and mice to study epilepsy or other neurological disorders.

Here, we introduce a novel technique designed to record electroencephalography (EEG) in freely moving neonatal epileptic pups and describe its procedures, features, and applications. This method allows one to record EEG for more than 1 week.

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such ...

Using Brain Activation (nir-HEG/Q-EEG) and Execution Measures (CPTs) in a ADHD Assessment Protocol

Attention Deficit Hyperactivity Disorder (ADHD) is a problem that impacts academic performance and has serious consequences that result in difficulties in scholastic, social and familial contexts. One of the most common problems in the identification of this disorder relates to the apparent over diagnosis of the disorder due to the absence of global protocols for assessment. The research group of School Learning, Difficulties and Academic Performance (ADIR) from the University of Oviedo, has developed a complete protocol that suggests the existence of certain patterns of cortical activation and executive control for identifying ADHD more objectively. This protocol takes into consideration some of the hypothetical determinants of ADHD, including the relationship between activation of selected areas of the brain, and differences in performance on various aspects of executive functioning such as omissions, commissions or response times, using innovative tools of Continuous Performance Testing (based on Virtual Reality CPT and Traditional CPT) and brain activation measures (two different tools, based on Hemoencephalography- nirHEG; and Quantified Electroencephalography --Q-EEG, respectively). This model of assessment aims to provide an effective assessment of ADHD symptomatology in order to design an accurate intervention and make appropriate recommendations for parents and teachers.

Using Brain Activation nir-HEG/Q-EEG and Execution Measures CPTs in a ADHD Assessment Protocol

This work presents a new protocol for the assessment of Attention Deficit Hyperactivity Disorder (ADHD) by providing a more objective diagnostic procedure for this developmental disorder based on the use of innovative tools. It also analyzes the relationship between activation measures and executive function measures.

Attention Deficit Hyperactivity Disorder (ADHD) is a problem that impacts academic performance and has serious consequences that result in difficulties in ...

Using Neuron Spiking Activity to Trigger Closed-Loop Stimuli in Neurophysiological Experiments

Closed-loop neurophysiological systems use patterns of neuronal activity to trigger stimuli, which in turn affect brain activity. Such closed-loop systems are already found in clinical applications, and are important tools for basic brain research. A particularly interesting recent development is the integration of closed-loop approaches with optogenetics, such that specific patterns of neuronal activity can trigger optical stimulation of selected neuronal groups. However, setting up an electrophysiological system for closed-loop experiments can be difficult. Here, a ready-to-apply Matlab code is provided for triggering stimuli based on the activity of single or multiple neurons. This sample code can be easily modified based on individual needs. For instance, it shows how to trigger sound stimuli and how to change it to trigger an external device connected to a PC serial port. The presented protocol is designed to work with a popular neuronal recording system for animal studies (Neuralynx). The implementation of closed-loop stimulation is demonstrated in an awake rat.

This protocol demonstrates how to use an electrophysiological system for closed-loop stimulation triggered by neuronal activity patterns. Sample Matlab code that can be easily modified for different stimulation devices is also provided.

Closed-loop neurophysiological systems use patterns of neuronal activity to trigger stimuli, which in turn affect brain activity. Such closed-loop systems ...

A Ferret Model of Inflammation-sensitized Late Preterm Hypoxic-ischemic Brain Injury

There is an ongoing need for clinically relevant models of perinatal infection and hypoxia-ischemia (HI) in which to test therapeutic interventions for infants with the neurological sequela of prematurity. Ferrets are ideal candidates for modeling the preterm human brain, as they are born lissencephalic and develop gyrencephalic brains postnatally. At birth, ferret brain development is similar to a 13 week human fetus, with postnatal-day (P) 17 kits considered to be equivalent to an infant at 32&#8211;36 weeks&#39; gestation. We describe an injury model in the P17 ferret, where lipopolysaccharide administration is followed by bilateral cerebral ischemia, hypoxia, and hyperoxia. This simulates the complex interaction of prolonged inflammation, ischemia, hypoxia, and oxidative stress experienced in a number of neonates who develop brain injury. Injured animals display a range of gross injury severity, with morphological changes in the brain including narrowing of multiple cortical gyri and associated sulci. Injured animals also show slowed reflex development, slower and more variable speed of locomotion in an automated catwalk, and decreased exploration in an open field. This model provides a platform in which to test putative therapies for infants with neonatal encephalopathy associated with inflammation and HI, study mechanisms of injury that affect cortical development, and investigate pathways that provide resilience in unaffected animals.

The method describes inflammation-sensitized hypoxic-ischemic and hyperoxic brain injury in the P17 ferret to model the complex interaction between prolonged inflammation and oxidative brain injury experienced in a number of late preterm infants.

There is an ongoing need for clinically relevant models of perinatal infection and hypoxia-ischemia (HI) in which to test therapeutic interventions for ...

Combining Behavior and EEG to Study the Effects of Mindfulness Meditation on Episodic Memory

Although there has been recent interest in how mindfulness meditation can affect episodic memory as well as brain structure and function, no study has examined the behavioral and neural effects of mindfulness meditation on episodic memory. Here we present a protocol that combines mindfulness meditation training, an episodic memory task, and EEG to examine how mindfulness meditation changes behavioral performance and the neural correlates of episodic memory. Subjects in a mindfulness meditation experimental group were compared to a waitlist control group. Subjects in the mindfulness meditation experimental group spent four weeks training and practicing mindfulness meditation. Mindfulness was measured before and after training using the Five Facet Mindfulness Questionnaire (FFMQ). Episodic memory was measured before and after training using a source recognition task. During the retrieval phase of the source recognition task, EEG was recorded. The results showed that mindfulness, source recognition behavioral performance, and EEG theta power in right frontal and left parietal channels increased following mindfulness meditation training. In addition, increases in mindfulness correlated with increases in theta power in right frontal channels. Therefore, results obtained from combining mindfulness meditation training, an episodic memory task, and EEG reveal the behavioral and neural effects of mindfulness meditation on episodic memory.

Here we present a protocol for combining mindfulness meditation training, an episodic memory task, and EEG to understand the behavioral and neural effects of mindfulness meditation on episodic memory.

Although there has been recent interest in how mindfulness meditation can affect episodic memory as well as brain structure and function, no study has ...

Modulation of the Neurophysiological Response to Fearful and Stressful Stimuli Through Repetitive Religious Chanting

In neuropsychological experiments, the late positive potential (LPP) is an event-related potential (ERP) component that reflects the level of one's emotional arousal. This study investigates whether repetitive religious chanting modulates the emotional response to fear- and stress-provoking stimuli, thus leading to a less responsive LPP. Twenty-one participants with at least one year of experience in the repetitive religious chanting of "Amitabha Buddha" were recruited. A 128-channel electroencephalography (EEG) system was used to collect EEG data. The participants were instructed to view negative or neutral pictures selected from the International Affective Picture System (IAPS) under three conditions: repetitive religious chanting, repetitive nonreligious chanting, and no chanting. The results demonstrated that viewing the negative fear- and stress-provoking pictures induced larger LPPs in the participants than viewing neutral pictures under the no-chanting and nonreligious chanting conditions. However, this increased LPP largely disappeared under repetitive religious chanting conditions. The findings indicate that repetitive religious chanting may effectively alleviate the neurophysiological response to fearful or stressful situations for practitioners.

The present event-related potential (ERP) study provides a unique protocol for investigating how religious chanting can modulate negative emotions. The results demonstrate that the late positive potential (LPP) is a robust neurophysiological response to negative emotional stimuli and can be effectively modulated by repetitive religious chanting.

In neuropsychological experiments, the late positive potential (LPP) is an event-related potential (ERP) component that reflects the level of one's ...

Automated Multimodal Stimulation and Simultaneous Neuronal Recording from Multiple Small Organisms

Fluorescent genetically encoded calcium indicators have contributed greatly to our understanding of neural dynamics from the level of individual neurons to entire brain circuits. However, neural responses may vary due to prior experience, internal states, or stochastic factors, thus generating the need for methods that can assess neural function across many individuals at once. Whereas most recording techniques examine a single animal at a time, we describe the use of wide-field microscopy to scale up neuronal recordings to dozens of Caenorhabditis elegans or other sub-millimeter-scale organisms at once. Open-source hardware and software allow great flexibility in programming fully automated experiments that control the intensity and timing of various stimulus types, including chemical, optical, mechanical, thermal, and electromagnetic stimuli. In particular, microfluidic flow devices provide precise, repeatable, and quantitative control of chemosensory stimuli with sub-second time resolution. The NeuroTracker semi-automated data analysis pipeline then extracts individual and population-wide neural responses to uncover functional changes in neural excitability and dynamics. This paper presents examples of measuring neuronal adaptation, temporal inhibition, and stimulus crosstalk. These techniques increase the precision and repeatability of stimulation, allow the exploration of population variability, and are generalizable to other dynamic fluorescent signals in small biosystems from cells and organoids to whole organisms and plants.

We present a method for the flexible chemical and multimodal stimulation and recording of simultaneous neural activity from many Caenorhabditis elegans worms. This method uses microfluidics, open-source hardware and software, and supervised automated data analysis to enable the measurement of neuronal phenomena such as adaptation, temporal inhibition, and stimulus crosstalk.

Fluorescent genetically encoded calcium indicators have contributed greatly to our understanding of neural dynamics from the level of individual neurons ...

OCT-Based Multimodal Imaging of Human Donor Eyes for AMD Research

Ex Vivo OCT-Based Multimodal Imaging of Human Donor Eyes for Research into Age-Related Macular Degeneration

A progression sequence for age-related macular degeneration (AMD) learned from optical coherence tomography (OCT)-based multimodal (MMI) clinical imaging could add prognostic value to laboratory findings. In this work, ex vivo OCT and MMI were applied to human donor eyes prior to retinal tissue sectioning. The eyes were recovered from non-diabetic white donors aged &#8805;80 years old, with a death-to-preservation time (DtoP) of &#8804;6 h. The globes were recovered on-site, scored with an 18 mm trephine to facilitate cornea removal, and immersed in buffered 4% paraformaldehyde. Color fundus images were acquired after anterior segment removal with a dissecting scope and an SLR camera using trans-, epi-, and flash illumination at three magnifications. The globes were placed in a buffer within a custom-designed chamber with a 60 diopter lens. They were imaged with spectral domain OCT (30&#176; macula cube, 30 &#181;m spacing, averaging = 25), near-infrared reflectance, 488 nm autofluorescence, and 787 nm autofluorescence. The AMD eyes showed a change in the retinal pigment epithelium (RPE), with drusen or subretinal drusenoid deposits (SDDs), with or without neovascularization, and without evidence of other causes. Between June 2016 and September 2017, 94 right eyes and 90 left eyes were recovered (DtoP: 3.9 &plusmn; 1.0 h). Of the 184 eyes, 40.2% had AMD, including early intermediate (22.8%), atrophic (7.6%), and neovascular (9.8%) AMD, and 39.7% had unremarkable maculas. Drusen, SDDs, hyper-reflective foci, atrophy, and fibrovascular scars were identified using OCT. Artifacts included tissue opacification, detachments (bacillary, retinal, RPE, choroidal), foveal cystic change, an undulating RPE, and mechanical damage. To guide the cryo-sectioning, OCT volumes were used to find the fovea and optic nerve head landmarks and specific pathologies. The ex vivo volumes were registered with the in vivo volumes by selecting the reference function for eye tracking. The ex vivo visibility of the pathology seen in vivo depends on the preservation quality. Within 16 months, 75 rapid DtoP donor eyes at all stages of AMD were recovered and staged using clinical MMI methods.

Author Spotlight: Ex Vivo OCT-Based Multimodal Imaging of Human Donor Eyes for Research into Age-Related Macular Degeneration  

Laboratory assays can leverage prognostic value from the longitudinal optical coherence tomography (OCT)-based multimodal imaging of age-related macular degeneration (AMD). Human donor eyes with and without AMD are imaged using OCT, color, near-infrared reflectance scanning laser ophthalmoscopy, and autofluorescence at two excitation wavelengths prior to tissue sectioning.

A progression sequence for age-related macular degeneration (AMD) learned from optical coherence tomography (OCT)-based multimodal (MMI) clinical imaging ...