Glass Microelectrodes Research Articles

Scanning glass microelectrode technique for investigating temperature-dependent electrical properties

Abstract Recent progresses in ionic current analyses related to micro- and nano-object sensing, electrochemical sensors, and liquid pollution monitoring have attracted significant attention. Micro- and nanoscale sensors with high spatial resolution and high signal-to-noise ratios are also effective for obtaining detailed understanding of ion transport phenomena. We have developed a glass microelectrode technique for measuring the electrical potential distribution by scanning through liquids. It enables us to directly evaluate electrical properties with a spatial resolution equal to the glass tip diameter, which is less than 1 μm. Herein, we optimize the channel and cell structures for the analysis of temperature-dependent properties, which allows us to measure the temperature dependence of conductivity and viscosity in the range of 303--333 K based on the Stokes--Einstein relation. The proposed method, which directly measures the spatial distribution of electrical potential, is suitable for analyzing conductivity, viscosity, and concentration without preprocessing calibration.

Fabrication of Microelectrodes, Suction Electrodes, and Focal Electrodes for Electrophysiological Recording in Drosophila.

Electrophysiological recording is a group of techniques used to record electrical field potentials generated by cells. These techniques rely on several types of electrodes, which can be manufactured in the laboratory. In intracellular recording, glass microelectrodes are used to pierce the cell membrane, and then to measure the electrical potential difference between the inside and the outside of the cell. Another technique, called loose patch or focal recording, is similar to intracellular recording but the electrode tip does not pierce into the cell membrane. Rather, the electrode tip is placed near a nerve or the postsynaptic side of the neuromuscular junction (NMJ) to record extracellular changes in local potentials. A third technique involves a suction electrode, which is used to draw part of the motor nerve into the electrode so that electrical pulses can be applied to elicit action potentials of the nerve. Suction electrodes are specifically used to evoke synaptic potentials at the Drosophila larval NMJ. This protocol details some basic methods for manufacturing microelectrodes used for intracellular recording and two-electrode voltage-clamp and loose patch electrodes used for focal recording. In addition, a method is provided for manufacturing homemade suction electrodes used for nerve stimulation.

The response of retinal ganglion cells to optical defocused visual stimuli in mouse retinas

Myopia and astigmatism are two primary types of refractive errors characterized by inaccurate focusing images on the retina. This study aimed to investigate the response characteristics of Retinal Ganglion Cells (RGCs), represented by alpha (α) RGCs, when exposed to focused, simulated spherically defocused images and astigmatically defocused images projected onto mouse retinas.Negative pressure was applied to stretch the soma of RGC in vitro to simulate myopia using a 7–8 μm diameter glass microelectrode, resulting in a 5% increase in the cell's diameter. A custom-made device was utilized to project spherically (equal to ±10 and ± 20 D) and astigmatically (+6.00 D) defocused images onto the retinas. As a control for a deficient intact retinal circuit, αRGCs of connexin 36 knockout (Cx36 KO) mice were used.The response of αRGCs varied significantly in terms of spikes, excitatory postsynaptic currents (EPSCs) and capacitances under stretching conditions to mimic myopia. Significant differences in the amplitudes of EPSCs were observed in the majority of αRGCs when exposed to focused and spherically defocused images in normal and mechanically simulated myopic retinas. However, this difference was not observed in αRGCs of Cx36 KO mice. αRGCs demonstrated significant differences in response between focused and astigmatically defocused images. Once again, αRGCs of Cx36 KO mice did not display differences.αRGCs have the ability to detect focused, spherically, and astigmatically defocused images and exhibit differential responses ex vivo. Gap junction subunit Cx36 may play a crucial role in transmitting visual signals associated with developing and perceiving refractive errors.

PH Sensing Based on Ionic Current Rectification Using Triple-Barreled Glass Microelectrodes

pH Sensing Based on Ionic Current Rectification Using Triple-Barreled Glass Microelectrodes

Electrical resistance of brain tissue during terminal ischemia

Terminal ischemia is characterized by several electrophysiological processes, including hyperpolarization, spreading depolarization (tSD), and negative ultraslow potentials (NUP) [1]. tSD is knowin to induce depolarization of neuron membranes to roughly 0 mV, accompanied by the influx of positive ions into cells, which results in the displacement of extracellular fluids into neurons, leading to cell swelling. In addition to the movement of extracellular fluid, cerebrospinal fluid flows into the perivascular region of the penetrating vessels. These processes result in a reduction of extracellular space and brain edema [2], which can elevate the likelihood of patient mortality up to 80% [3]. The decrease in extracellular space may lead to an upsurge in the electrical resistance of the tissue, enabling it to function as an important indicator of the anatomical and functional state of the brain during traumatic injuries and hemorrhages [4]. Our goal was to assess the resistance of the extracellular space in the rat’s barrel cortex by measuring voltage step amplitudes caused by current injection between the V1 cortex and the tail vein of an animal. We used 16-channel probes with iridium electrode sites and glass microelectrodes containing Ag/AgCl conductors filled with NaCl solution. We used a metal Ag/AgCl electrode as a reference, implanted in the cerebellum. The experimental animals were euthanized via inhalation of isoflurane at a lethal concentration. Isoflurane-induced respiratory arrest led to the subsequent development of a sequence of electrophysiological processes that included hyperpolarization, terminal spreading depression, and negative ultraslow potential. The NUP persisted throughout the entirety of the recording period (30–90 minutes), with a significant increase in extracellular space resistance occurring during this time. The rise in resistance began concurrently with the end of respiration. The resistance increased by 40 [23–57]% (median [25th–75th percentile], p=0.002, n=11) after 30 minutes of breath cessation, and by 46 [(–15)–64]% (p=0.109, n=8) after 60 minutes. The resistance demonstrated equivalent changes in all cerebral cortex depths. Additionally, the NUP amplitude following respiratory arrest had a correlation with the increase of voltage step amplitudes at 30 minutes (R=–0.713, p=0.014). The resistances measured using signals from both Ag/AgCl- and Ir electrodes did not vary in any of the experiments conducted (p=1, n=5). Thus, the rise in electrical resistance in brain tissue was shown to initiate at the point of respiratory arrest in the animal and progresses alongside the terminal processes in the cerebral cortex. This correlation is specifically linked to the fluctuations in the negative ultraslow potential. In tDS and NUP, a significant increase in tissue edema occurs due to the movement of water from the extracellular spaces into the cells, which leads to a decrease in extracellular space volume and an increase in resistance. The process can last for several minutes and is characterized by a continuous increase in voltage step amplitudes. These results suggest that tissue resistance increases during focal ischemia, as SD and NUP are also present during the formation of the ischemic focus [5].

Placement of Extracranial Stimulating Electrodes and Measurement of Cerebral Blood Flow and Intracranial Electrical Fields in Anesthetized Mice.

The detection of cerebral blood flow (CBF) responses to various forms of neuronal activation is critical for understanding dynamic brain function and variations in the substrate supply to the brain. This paper describes a protocol for measuring CBF responses to transcranial alternating current stimulation (tACS). Dose-response curves are estimated both from the CBF change occurring with tACS (mA) and from the intracranial electric field (mV/mm). We estimate the intracranial electrical field based on the different amplitudes measured by glass microelectrodes within each side of the brain. In this paper, we describe the experimental setup, which involves using either bilateral laser Doppler (LD) probes or laser speckle imaging (LSI) to measure the CBF; as a result, this setup requires anesthesia for the electrode placement and stability. We present a correlation between the CBF response and the current as a function of age, showing a significantly larger response at higher currents (1.5 mA and 2.0 mA) in young control animals (12-14 weeks) compared to older animals (28-32 weeks) (p < 0.005 difference). We also demonstrate a significant CBF response at electrical field strengths <5 mV/mm, which is an important consideration for eventual human studies. These CBF responses are also strongly influenced by the use of anesthesia compared to awake animals, the respiration control (i.e., intubated vs. spontaneous breathing), systemic factors (i.e., CO2), and local conduction within the blood vessels, which is mediated by pericytes and endothelial cells. Likewise, more detailed imaging/recording techniques may limit the field size from the entire brain to only a small region. We describe the use of extracranial electrodes for applying tACS stimulation, including both homemade and commercial electrode designs for rodents, the concurrent measurement of the CBF and intracranial electrical field using bilateral glass DC recording electrodes, and the imaging approaches. We are currently applying these techniques to implement a closed-loop format for augmenting the CBF in animal models of Alzheimer's disease and stroke.

Study on the central neural pathway and the relationship between the heart and small intestine via a dual neural tracer.

Despite very different functions, studies increasingly report that there may be a potential central nervous anatomical connection between the heart and the small intestine. In this study, the central nervous anatomical relationship between the heart and small intestine was studied via a viral tracer. Pseudorabies virus (PRV) syngeneic strains with different fluorescent reporter genes (eGFP or mRFP) were microinjected into the heart walls and small intestinal walls of male C57BL/6J using glass microelectrode. The results showed that the co-labeled nuclei in the brain were lateral periaqueductal gray (LPAG) and ventrolateral periaqueductal gray (VLPAG) in the midbrain, mesencephalic trigeminal nucleus (Me5), and motor trigeminal nucleus anterior digastric Part (5Adi) in the pons. The co-labeled sites in the spinal cord were intermediolateral column (IML) in the second thoracic vertebra, IML and lamina 7 of the spinal gray (7SP) in the third thoracic vertebra, and IML in the fourth thoracic vertebra. Our data show that there is a neuroanatomical connection between the small intestine and the heart in the central nervous system (CNS). Neuroanatomical integration of the heart and small intestine may provide a basis for revealing the physiological and pathological interactions between the circulatory and digestive systems. The interactions may be mediated more effectively through sympathetic nerves.

Abstract 12951: Electrical and Calcium-Dependent Mechanisms That Contribute to Increase the Risk for Atrial Fibrillation in Obesity

Obesity (OB) is an independent predictor for atrial fibrillation (AF), but mechanisms by which OB increases AF risk remain poorly characterized. We hypothesized that OB modulates AF risk through electrical remodeling and impaired Ca 2+ dynamics that can be reversed after weight loss (WL). Using transesophageal rapid pacing we characterized AF inducibility and burden in lean, OB, and OB mice that lost 20% of body weight after modified vertical sleeve gastrectomy. Local field fluorescence microscopy was used to simultaneously record action potentials (APs) (di-8-ANEPPS or glass microelectrodes) and Ca 2+ transients (CaT) (Rhod-2) in Langendorff heart preparations. We evaluated AP duration (APD) dependency of frequency from 4 to 14 Hz (1 Hz incremental steps). AP parameters analyzed included APD30, APD50, APD90 and spontaneous arrhythmogenic events. We assessed Ca 2+ dynamics by studying CaT and fractional sarcoplasmic reticulum Ca 2+ release in the absence and presence of ryanodine (Ry) and thapsigargin (Tg). Since CaT Ry+Tg represents sarcolemmal Ca 2+ influx, we calculated the gain of Ca 2+ induced Ca 2+ release (CICR gain ) as the ratio between CaT in the absence and presence of Ry+Tg. Experiments were done in the presence of blebbistatin at 37° C. Mean±SE, t-test. After burst pacing, both AF prevalence and burden were higher in OB mice vs lean (94 % vs 47 %, 227±19 vs 20±1.2 s, n=16 mice/group, p&lt;0.0001). About 76 % of OB hearts showed afterdepolarizations and fibrillation vs no events in lean. APD50 and APD90 significantly decreased as stimulation frequency increased from 4 to 14 Hz in OB hearts vs lean (n=6 mice/group, p&lt;0.05). Peak CaT from OB hearts were significantly decreased compared to peak CaT from lean (0.3±0.05 vs. 0.8±0.06, n=9, p&lt;0.001). CICR gain in OB hearts was also significantly compromised compared to lean (5.2±1 vs 23.3±1.4, n=7 mice/group, p&lt;0.0001). After WL, OB mice showed a decreased AF burden (21.7±8 s), increased APD, increased peak CaT (0.8±0.15) and increased CICR gain (20±2.3) back to lean levels (n=8, p&lt;0.001). In conclusion, shortened APs, spontaneous afterdepolarizations and impaired CICR significantly contribute to the development of AF in OB. WL reversed the electrical phenotype and the impaired Ca 2+ dynamics and prevented AF.

Local Electric Field and Electrical Conductivity Analysis Using a Glass Microelectrode.

Transport phenomena in microfluidic chips are induced by electric fields and electrolyte concentrations. Liquid flows are often affected by ionic currents driven by electric fields in narrow channels, which are applied in microelectromechanical systems, microreactors, lab-on-a-chip, and so forth. Even though numerical studies to evaluate those local fields have been reported, measurement methods seem to be under construction. To deeply understand the dynamics of ions at the microscale, measurement techniques are necessary to be developed. In this study, we propose a novel method to directly measure electrical potential differences in liquids, local electric fields, and electrical conductivities, using a glass microelectrode. Scanning an electrolyte solution, for example, KCl solutions, with a 1 μm tip under constant ionic current conditions, a potential difference in liquids is locally measured with a micrometer-scale resolution. The conductivity of KCl solutions ranging from 0.56 to 100 mM is evaluated from electric fields locally measured, and errors are within 5% compared with the reference values. It is found that the present method enables us to directly measure local electric fields under constant current and that the electrical conductivity is quantitatively evaluated. Furthermore, it is suggested that the present method is available for various electrical analyses without calibration procedures before measurements.

A 146-dB-Ohm Gain 14-pARMS Noise Patch-Clamp Amplifier for Whole-Cell Ion Current Detection

The membrane clamp technique is an important tool to reflect the electrophysiological characteristics of cells by recording the ionic currents of cellular channels. Embedded systems work in systems designed for specific user groups and are used to implement specific functions. The membrane clamp amplifier built with embedded technology has the advantages of miniaturization, specialization, low power consumption, high integration, high resource utilization, and a long life cycle, which can avoid the inconvenience to developers and experimenters due to the update of general-purpose computer software and hardware. In this paper, a patch-clamp amplifier (PCA) based on a transimpedance amplifier (TIA) is proposed, which includes a glass microelectrode series resistance/capacitance compensation circuit and feedback resistor parasitic capacitance compensation. The prototype is designed using 180 nm CMOS technology and occupies an area of 720 μ m × 630 μ m . The prototype achieves a transimpedance gain of 145.6 dB-Ohm, a -3 dB bandwidth greater than 15 kHz, and an input reference noise current of 13.98 pARMS. Experimental results show that the proposed PCA is capable of compensating up to 30 pF of electrode capacitance and up to 10 MΩ for 86% of the series resistance, respectively. In addition, when the feedback resistor parasitic capacitance compensation circuit is enabled, the overshoot phenomenon disappears, overcoming the shortcomings of the conventional diaphragm clamp amplifier current clamp and maintaining the original key performance.

SGLT2 Inhibitor Empagliflozin Modulates Ion Channels in Adult Zebrafish Heart.

Empagliflozin, an inhibitor of sodium-glucose co-transporter 2 (iSGLT2), improves cardiovascular outcomes in patients with and without diabetes and possesses an antiarrhythmic activity. However, the mechanisms of these protective effects have not been fully elucidated. This study aimed to explore the impact of empagliflozin on ion channel activity and electrophysiological characteristics in the ventricular myocardium. The main cardiac ionic currents (INa, ICaL, ICaT, IKr, IKs) and action potentials (APs) were studied in zebrafish. Whole-cell currents were measured using the patch clamp method in the isolated ventricular cardiomyocytes. The conventional sharp glass microelectrode technique was applied for the recording of APs from the ventricular myocardium of the excised heart. Empagliflozin pretreatment compared to the control group enhanced potassium IKr step current density in the range of testing potentials from 0 to +30 mV, IKr tail current density in the range of testing potentials from +10 to +70 mV, and IKs current density in the range of testing potentials from −10 to +20 mV. Moreover, in the ventricular myocardium, empagliflozin pretreatment shortened AP duration APD as shown by reduced APD50 and APD90. Empagliflozin had no influence on sodium (INa) and L- and T-type calcium currents (ICaL and ICaT) in zebrafish ventricular cardiomyocytes. Thus, we conclude that empagliflozin increases the rapid and slow components of delayed rectifier K+ current (IKr and IKs). This mechanism could be favorable for cardiac protection.

Neural mechanism for modulation of auditory response of the striatum by locomotion

To explore whether the characteristic responses to sound stimulations of the auditory neurons in the striatum is regulated in different behavioral states. The auditory neurons in the striatum of awake C57BL/6J mice were selected for this study. We recorded the auditory response of the striatum to noises over a long period of time by building a synchronous in vivo electrophysiological and locomotion recording system and using glass microelectrode attachment recording. By analyzing the running speed of the mice, the behavioral states of the mice were divided into the quiet state and the active state, and the spontaneous activity and evoked responses of the auditory neurons in the striatum were analyzed in these two states. Compared with those recorded in the quiet state, the spontaneous activity of the auditory neurons in the striatum of the mice increased significantly (37.06±12.02 vs 18.51±10.91, P < 0.001) while the auditory response of the neurons decreased significantly (noise intensity=60 dB, 3.45±2.99 vs 3.04±2.76, P < 0.001) in the active state. Locomotion has a significant inhibitory effect on the auditory response of the striatum, which may importantly contribute to the decline of sound information recognition ability in the active state.

Characterizing the Luminal Microenvironment of Human Organoids for Studies of Gastric pH Regulation

The stomach is notorious for having the steepest proton gradient in the body, which pathogenic Helicobacter pylori must traverse to infect the epithelium. This gradient across the 200‐300 μm‐thick gastric mucus layer enables the stomach to maintain a near‐neutral pH at the epithelial surface and a strongly acidic environment in the stomach lumen. In this study, we use human gastric organoids to study the maintenance and regulation of this pH gradient. Human gastric organoids are 3D cellular constructs derived from patient biopsies that mimic the microanatomy and physiology of the stomach, as they contain both mucus and gastric acid. Organoids have been used to investigate H. pylori‐induced gastritis, peptic ulcers, and stomach cancer, which claims 700,000 lives each year. Gastric mucus is critical for protection against H. pylori infection, which is the leading cause of gastric cancer. It is presently unclear how the gastric mucosal pH gradient is maintained, and how closely organoid models can mimic this microenvironment. First, we detected mucus within human gastric organoids using histological staining and bead microrheology. We also used a FITC‐conjugated lectin to visualize the mucosal matrix inside of the organoids. These data were supported by our observation that a clear viscous material was produced on the apical side of two‐dimensional, organoid‐derived cell monolayers. Additionally, we developed a novel method for the measurement of intraluminal pH of the gastric organoids. Using a micromanipulator and stereomicroscope to maneuver a glass microelectrode into the organoid lumen, we measured and manipulated luminal pH using histamine stimulation. We determined that pH remains consistent across organoid lines, and that the intraluminal space is more acidic than the surrounding environment. In ongoing studies, we will develop the spatial resolution of our organoid microprofiling technique to fully characterize the proton gradient within the mucus‐filled lumens gastric organoids. Our studies are the first to utilize pH microsensors in these models, and will lead to improved molecular and biophysical understanding of gastric physiology and potential new treatment approaches for disease conditions (such as peptic ulcers and gastric cancer) that involve dysregulated gastric acid secretion.

Capturing individual charged microparticles and cells with microelectrodes in a circular microfluidic channel

Capture and, subsequently, in situ characterization of microparticles generally require complex fabrication processes and sophisticated techniques in microfluidic devices. In this paper, we present a simple and inexpensive technique for capturing and applying chemical agents to individual charged microparticles with the same glass microelectrode in a circular microfluidic channel. The microfluidic device features a thin poly-dimethyl-siloxane (PDMS) layer (100 µm, thickness) above the circular channel (diameter, 50 µm) so that the tip of the glass microelectrode can enter into the channel by penetrating through the thin PDMS layer. When an electric field is applied across the tip of the microelectrodes, the tip can capture charged microparticles or living cells that are flowing through the channel. Meanwhile, the same glass microelectrode can be used to apply chemical agents to the captured particles or living cells by filling the solution of the chemical agent into its tip. This simple design method offered simple fabrication without the need for any specialized equipment for the manipulation of charged particles or living cells in controlled microenvironments.

Mechanisms of cancellation by pain of genetically determined inhibition of a malignant tumor growth in experiment

Aims are to study the nature of the processes of carcinogenesis of experimental B16/F10 melanoma in uPA gene knockout mice modified by chronic neurogenic pain and investigate some electrophysiological mechanisms of melanoma development. Materials and methods. We used 48 C57BL/6-PlautmI. IBug-ThisPlau6FDhu/GFDhu mice of both genders with urokinase gene knockout and 102 C57BL/6 mice of both genders with the normal genotype. Chronic neurogenic pain (CNP) was produced due to bilateral ligation of the sciatic nerve. Against the above background, all animals were transplanted with B16/F10 melanoma. To study the mechanism of CNP, studies of the intracellular electrophysiological activity of neurons of the central nervous system of the snail Helix pomatia in the body in vivo were carried out. CNP was reproduced by dosed pressing of four main nerves with Fresnel hairs that with time turned into increasing pain. Membrane potential (MP), action potential (AP) and firing rate (FR) parameters of intracellular bio-potentials of the command neuron RPaG3, continuously recorded using an ultrathin glass microelectrode for 4-5 days, were analyzed. Results. It was detected that an activation of cancerogenesis during the modification of the progression of experimental B16/ F10 melanoma in C57BL/6-PlautmI.IBug-ThisPlau6FDhu/GFDhu mice with uPA gene knockout using CNP is accompanied by a 2-fold acceleration in the time of tumor production, stimulation of the growth of the primary tumor nodes from 1.05±0.08 cm3 to 9.50±0.98 cm3 (p&lt;0.001) and multiple metastasizing to the lungs, a reduction in the life span from 34.67±0.67 to 21.33±2.19 days (p&lt;0.001) &lt;0.05) in the genetically modified mice, by changing some gender-specific characteristics of the progression of the malignant process. The neuropathic nature of pain resulting from command neuron compression or ligation of the sciatic nerves is essentially identical to the implementation of genetic programs responsible for the control of life and death, reproducing events in carcinogenesis with the progression of a malignant tumor. Conclusion. The initiation and chronization of pain at the local level of the nervous system can lead to generalization of the pain syndrome and contribute to the cancellation of genetically predetermined programs of carcinogenesis.

Development of direct analysis method of conductivity by local electric field measurement

Recently, micro- and nanofluidics devices such as MEMS/NEMS and μ-TAS have attracted much attention. However, several unknowns have been remained in micro- and nanoscale transport phenomena unlike microscopic scale. In order to deepen understanding of such micro- and nanoscale phenomena, it is essential to develop a novel technique to measuer the small spaces. In this study, a localized and direct measurement method of electric fields is established by using a test section with a charcteristic length of 1 mm and a glass microelectrode. Using a KCl standard solution ranging from 0.56 to 100 mmol/L as a sample, the conductivity is successfully evaluated with a measurement error of less than 15%.

Impacts of a high-fat diet on the metabolic profile and the phenotype of atrial myocardium in mice.

Obesity, diabetes, and metabolic syndromes are risk factors of atrial fibrillation (AF). We tested the hypothesis that metabolic disorders have a direct impact on the atria favouring the formation of the substrate of AF. Untargeted metabolomic and lipidomic analysis was used to investigate the consequences of a prolonged high-fat diet (HFD) on mouse atria. Atrial properties were characterized by measuring mitochondria respiration in saponin-permeabilized trabeculae, by recording action potential (AP) with glass microelectrodes in trabeculae and ionic currents in myocytes using the perforated configuration of patch clamp technique and by several immuno-histological and biochemical approaches. After 16 weeks of HFD, obesogenic mice showed a vulnerability to AF. The atrial myocardium acquired an adipogenic and inflammatory phenotypes. Metabolomic and lipidomic analysis revealed a profound transformation of atrial energy metabolism with a predominance of long-chain lipid accumulation and beta-oxidation activation in the obese mice. Mitochondria respiration showed an increased use of palmitoyl-CoA as energy substrate. APs were short duration and sensitive to the K-ATP-dependent channel inhibitor, whereas K-ATP current was enhanced in isolated atrial myocytes of obese mouse. HFD transforms energy metabolism, causes fat accumulation, and induces electrical remodelling of the atrial myocardium of mice that become vulnerable to AF.

The role of M3 receptors in regulation of electrical activity deteriorates in the rat heart during ageing.

Ageing is a complex process which affects all systems of the organism and therefore changes the environment where the heart is working. In this study we demonstrate the ageing-related changes in the mechanisms of parasympathetic regulation of mammalian heart. Electrophysiological effects produced by selective activation of M3-cholinoreceptors were compared in isolated cardiac preparations from young adult (4 months), adult (1 year) and ageing (2 years) rats using sharp glass microelectrode technique. M3-receptors were activated with muscarinic agonist pilocarpine (10-5M) in the presence of selective M2 antagonist AQ-RA741 (10-7M). In atrial and ventricular myocardium from young rats M3 stimulation induced shortening of action potentials(APs), while no significant effect was observed in both elder groups. The main mechanism of M3-induced AP shortening is inhibition of L-type Ca2+ current, estimated using whole-cell patch-clamp. It was negligible in atrial myocytes from ageing animals in comparison with young rats. The loss of sensitivity to stimulation of M3-receptors is due to decrease in M3 gene expression, shown by RT-PCR both in atrial and ventricular samples from ageing rats. Thus, in ageing rat heart M3-receptors are down-regulated and not involved in regulation of electrical activity.

Carbon fiber electrodes for intracellular recording and stimulation

Objective. To understand neural circuit dynamics, it is critical to manipulate and record many individual neurons. Traditional recording methods, such as glass microelectrodes, can only control a small number of neurons. More recently, devices with high electrode density have been developed, but few of them can be used for intracellular recording or stimulation in intact nervous systems. Carbon fiber electrodes (CFEs) are 8 µm-diameter electrodes that can be assembled into dense arrays (pitches ⩾ 80 µm). They have good signal-to-noise ratios (SNRs) and provide stable extracellular recordings both acutely and chronically in neural tissue in vivo (e.g. rat motor cortex). The small fiber size suggests that arrays could be used for intracellular stimulation. Approach. We tested CFEs for intracellular stimulation using the large identified and electrically compact neurons of the marine mollusk Aplysia californica. Neuron cell bodies in Aplysia range from 30 µm to over 250 µm. We compared the efficacy of CFEs to glass microelectrodes by impaling the same neuron’s cell body with both electrodes and connecting them to a DC coupled amplifier. Main results. We observed that intracellular waveforms were essentially identical, but the amplitude and SNR in the CFE were lower than in the glass microelectrode. CFE arrays could record from 3 to 8 neurons simultaneously for many hours, and many of these recordings were intracellular, as shown by simultaneous glass microelectrode recordings. CFEs coated with platinum-iridium could stimulate and had stable impedances over many hours. CFEs not within neurons could record local extracellular activity. Despite the lower SNR, the CFEs could record synaptic potentials. CFEs were less sensitive to mechanical perturbations than glass microelectrodes. Significance. The ability to do stable multi-channel recording while stimulating and recording intracellularly make CFEs a powerful new technology for studying neural circuit dynamics.

Evolution of auditory response signal-to-noise ratio in ascending auditory pathways

To explore the characteristics of the evolution of auditory response signal-to-noise ratio at all levels of the ascending auditory pathway, its modulation by different brain states in different brain regions, and its potential value as an effective indicator for encoding sound characteristics. Eighty C57BL/6J awake mice were used for recording the best frequency auditory response of the neurons in the inferior colliculus (n=20), medial geniculate body (n=20), and primary auditory cortex using a glass microelectrode. The probability density of spontaneous and evoked firing of the neurons was calculated to establish a distribution model of spontaneous and evoked firing, and the evolution of the auditory response signal-to-noise ratio was statistically analyzed. The changes in spontaneous and evoked firing of the neurons and the auditory response signal-to-noise ratio in different brain regions were analyzed at rest and during running. In different brain regions in the ascending auditory pathway, the spontaneous firing of the neurons all showed a Poisson distribution, and the evoked firing showed a lognormal distribution. The auditory response signal-to-noise ratio was significantly greater in the inferior colliculus than in the medial geniculate body and auditory cortex (P < 0.001). The auditory response signal-to-noise ratio in the 3 brain regions remained stable irrespective of the states of motion (P>0.05). Auditory response signal-to-noise ratio may serve as an effective indicator of encoding sound characteristics.