Components Of Action Potential Research Articles

The role of spinal cord neuroanatomy in the variances of epidural spinal recordings

BackgroundSpinal cord stimulation (SCS) has demonstrated multiple benefits in treating chronic pain and other clinical disorders related to sensorimotor dysfunctions. However, the underlying mechanisms are still not fully understood, including how electrode placement in relation to the spinal cord neuroanatomy influences epidural spinal recordings (ESRs). To characterize this relationship, this study utilized stimulation applied at various anatomical sections of the spinal column, including at levels of the intervertebral disc and regions correlating to the dorsal root entry zone.MethodTwo electrode arrays were surgically implanted into the dorsal epidural space of the swine. The stimulation leads were positioned such that the caudal-most electrode contact was at the level of a thoracic intervertebral segment. Intraoperative cone beam computed tomography (CBCT) images were utilized to precisely determine the location of the epidural leads relative to the spinal column. High-resolution microCT imaging and 3D-model reconstructions of the explanted spinal cord illustrated precise positioning and dimensions of the epidural leads in relation to the surrounding neuroanatomy, including the spinal rootlets of the dorsal and ventral columns of the spinal cord. In a separate swine cohort, implanted epidural leads were used for SCS and recording evoked ESRs.ResultsReconstructed 3D-models of the swine spinal cord with epidural lead implants demonstrated considerable distinctions in the dimensions of a single electrode contact on a standard industry epidural stimulation lead compared to dorsal rootlets at the dorsal root entry zone (DREZ). At the intervertebral segment, it was observed that a single electrode contact may cover 20-25% of the DREZ if positioned laterally. Electrode contacts were estimated to be ~0.75 mm from the margins of the DREZ when placed at the midline. Furthermore, ventral rootlets were observed to travel in proximity and parallel to dorsal rootlets at this level prior to separation into their respective sides of the spinal cord. Cathodic stimulation at the level of the intervertebral disc, compared to an ‘off-disc’ stimulation (7 mm rostral), demonstrated considerable variations in the features of recorded ESRs, such as amplitude and shape, and evoked unintended motor activation at lower stimulation thresholds. This substantial change may be due to the influence of nearby ventral roots. To further illustrate the influence of rootlet activation vs. dorsal column activation, the stimulation lead was displaced laterally at ~2.88 mm from the midline, resulting in variances in both evoked compound action potential (ECAP) components and electromyography (EMG) components in ESRs at lower stimulation thresholds.ConclusionThe results of this study suggest that the ECAP and EMG components of recorded ESRs can vary depending on small differences in the location of the stimulating electrodes within the spinal anatomy, such as at the level of the intervertebral segment. Furthermore, the effects of sub-centimeter lateral displacement of the stimulation lead from the midline, leading to significant changes in electrophysiological metrics. The results of this pilot study reveal the importance of the small displacement of the electrodes that can cause significant changes to evoked responses SCS. These results may provide further valuable insights into the underlying mechanisms and assist in optimizing future SCS-related applications.

Short-term plasticity as 'energetic memory' of ion channel components of action potential.

Information transfer in the nervous system is traditionally understood by the transmission of action potentials along neuronal dendrites, with ion channels in the membrane as the basic unit operator for their creation and propagation. We present here a new model for the multiphysics behaviour of ion channels and the action potential dynamics in nervous and other signal-transmitting systems. This model is based on the long-term suppression of an action potential as a response to mechanical input. While other models focus on electrical aspects of the action potential, an increasing body of experiments highlights its electro-mechanical nature and points in particular towards an alteration of the action potential when subjected to a mechanical input. Here, we propose a new phenomenological framework able to capture the mechanical aspect of ion channel dynamics and the resulting effect on the overall electrophysiology of the membrane. The model is introduced here through a set of coupled differential equations that describe the system while agreeing with the general findings of the experiments that support an electro-mechanical model. It also confirms that transient quasi-static mechanical loads reversibly affect the amplitude and rate of change of neuronal action potentials, which are smaller and slower under indentation loading conditions. Changes after the loading release are also reversible, albeit on a different time scale.

Photobiomodulation for pain relief: Model-based estimates of effective doses of light at the neural target

IntroductionPhotobiomodulation (PBM) has been studied since the 1960s as a clinical tool. More recently, PBM has been observed to reduce compound action potential components and hypersensitivities associated with neuropathic pains. However, no definitive description of efficacious light parameters has been determined. Some reasons may be that previous meta-analyses and reviews have focused on emitter output rather than the light at the target tissue and have included data sets that are large but with notable variability (e.g., combining data from various disease etiologies, and data from PBM at various wavelengths). This fact has made it difficult to successfully define the range of effective parameters. MethodsIn this study, photon propagation software was used to estimate irradiance at a target nerve using several published data sets chosen for their narrow criteria to minimize variability. Utilizing these estimates, effective and ineffective light irradiances at the nerve of interest for wavelengths of 633 nm or 808–830 nm were examined and estimated. These estimates are focused on the amount of light required to achieve a reduction in pain or a surrogate measure via a hypothesized nerve block mechanism. ResultsAccounting for irradiance at the target nerve yielded a clear separation of PBM doses that achieved small-fiber nerve block from those that did not. For both the 633 nm group and the 808–830 group, the irradiance separation threshold followed a nonlinear path with respect to PBM application duration, where shorter durations required higher irradiances, and longer durations required lower irradiances. Using the same modeling methods, irradiance was estimated as a function of depth from a transcutaneous source (distance from skin surface) for emitter output power using small or large emitter sizes. ConclusionTaken together, the results of this study can be used to estimate effective PBM dosing schemes to achieve small-fiber inhibition for various anatomical scenarios.

ID: 213423 Changes in Evoked Responses During Epidural Spinal Cord Stimulation Generated by Minimal Movement of Leads

Lead migration is a frequent problem with Spinal Cord Stimulation (SCS) for chronic pain. The role of lead migration in efficacy of pain control is unclear. Using a large animal model, we investigated if a movement of stimulation lead could cause change in the evoked response, which could place impact on the evoked compound action potential (ECAP) component, seeking a way to notify and quantify lead migration during SCS.

Median-to-Ulnar Nerve Communication in Carpal Tunnel Syndrome: An Electrophysiological Study.

The median-to-ulnar communicating branch (MUC) is an asymptomatic variant of the upper limb innervation that can lead to interpretation errors in routine nerve conduction studies. The diagnosis of carpal tunnel syndrome (CTS) or ulnar nerve lesions can be complicated by the presence of MUC. In this study, we describe electrophysiological features of MUC in CTS patients presenting to our clinic. We enrolled MUB cases from consecutive CTS patients referred to our laboratory between the years 2014 and 2019. MUC was present in 53 limbs (36 patients) from the studied population. MUC was bilateral in 53% of patients. MUC type II was the most common subtype (74%), followed by types III and I; more coexisting MUC types were found in the majority of tested limbs. A positive correlation was demonstrated between the severity of CTS and the presence of positive onset, faster CV, or a double component of the compound muscle action potentials. We emphasize the importance of suspecting the presence of MUC in CTS in the presence of a positive onset or a double component in routine motor conduction studies.

Slow recovery from the inactivation of voltage-gated sodium channel Nav1.3 in mouse taste receptor cells.

Action potentials play an important role in neurotransmitter release in response to taste. Here, I have investigated voltage-gated Na+ channels, a primary component of action potentials, in respective cell types of mouse fungiform taste bud cells (TBCs) with in situ whole-cell clamping and single-cell RT-PCR techniques. The cell types of TBCs electrophysiologically examined were determined immunohistochemically using the type III inositol 1,4,5-triphoshate receptor as a type II cell marker and synaptosomal-associated protein 25 as a type III cell marker. I show that type II cells, type III cells, and TBCs not immunoreactive to these markers (likely type I cells) generate voltage-gated Na+ currents. The recovery following inactivation of these currents was well fitted with double exponential curves. The time constants in type III cells (~20ms and ~ 1s) were significantly slower than respective time constants in other cell types. RT-PCR analysis indicated the expression of Nav1.3, Nav1.5, Nav1.6, and β1 subunit mRNAs in TBCs. Pharmacological inhibition and single-cell RT-PCR studies demonstrated that type II and type III cells principally express tetrodotoxin (TTX)-sensitive Nav1.3 channels and that ~ 30% of type I cells express TTX-resistant Nav1.5 channels. The auxiliary β1 subunit that modulates gating kinetics was rarely detected in TBCs. As the β1 subunit co-expressed with an α subunit is known to accelerate the recovery from inactivation, it is likely that voltage-gated Na+ channels in TBCs may function without β subunits. Slow recovery from inactivation, especially in type III cells, may limit high-frequency firing in response to taste substances.

Increased Axon Initial Segment Length Results in Increased Na+ Currents in Spinal Motoneurones at Symptom Onset in the G127X SOD1 Mouse Model of Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease preferentially affecting motoneurones. Transgenic mouse models have been used to investigate the role of abnormal motoneurone excitability in this disease. Whilst an increased excitability has repeatedly been demonstrated in vitro in neonatal and embryonic preparations from SOD1 mouse models, the results from the only studies to record in vivo from spinal motoneurones in adult SOD1 models have produced conflicting findings. Deficits in repetitive firing have been reported in G93A SOD1(high copy number) mice but not in presymptomatic G127X SOD1 mice despite shorter motoneurone axon initial segments (AISs) in these mice. These discrepancies may be due to the earlier disease onset and prolonged disease progression in G93A SOD1 mice with recordings potentially performed at a later sub-clinical stage of the disease in this mouse. To test this, and to explore how the evolution of excitability changes with symptom onset we performed in vivo intracellular recording and AIS labelling in G127X SOD1 mice immediately after symptom onset. No reductions in repetitive firing were observed showing that this is not a common feature across all ALS models. Immunohistochemistry for the Na+ channel Nav1.6 showed that motoneurone AISs increase in length in G127X SOD1 mice at symptom onset. Consistent with this, the rate of rise of AIS components of antidromic action potentials were significantly faster confirming that this increase in length represents an increase in AIS Na+ channels occurring at symptom onset in this model.

Perfidious synaptic transmission in the guinea-pig auditory brainstem.

The presence of ‘giant’ synapses in the auditory brainstem is thought to be a specialization designed to encode temporal information to support perception of pitch, frequency, and sound-source localisation. These ‘giant’ synapses have been found in the ventral cochlear nucleus, the medial nucleus of the trapezoid body and the ventral nucleus of the lateral lemniscus. An interpretation of these synapses as simple relays has, however, been challenged by the observation in the gerbil that the action potential frequently fails in the ventral cochlear nucleus. Given the prominence of these synapses it is important to establish whether this phenomenon is unique to the gerbil or can be observed in other species. Here we examine the responses of units, thought to be the output of neurons in receipt of ‘giant’ synaptic endings, in the ventral cochlear nucleus and the medial nucleus of the trapezoid body in the guinea pig. We found that failure of the action-potential component, recorded from cells in the ventral cochlear nucleus, occurred in ~60% of spike waveforms when recording spontaneous activity. In the medial nucleus of the trapezoid body, we did not find evidence for action-potential failure. In the ventral cochlear nucleus action-potential failures transform the receptive field between input and output of bushy cells. Additionally, the action-potential failures result in “non-primary-like” temporal-adaptation patterns. This is important for computational models of the auditory system, which commonly assume the responses of ventral cochlear nucleus bushy cells are very similar to their “primary like” auditory-nerve-fibre inputs.

Thermal dependency of electrical excitability of gastrocnemius (skeletal) muscle in the desert-reptile, Uromastix hardwickii

This study deals with effects of seasonal temperature variations on muscular electrical excitability in the reptile Uromastix hardwickii. Freshly captured adult animals of both sexes were used, and the gastrocnemius (skeletal) muscles were dissected out. The muscle samples were digested with digestive fluid (pepsin and HCl), stirred, and the cells were allowed to settle out. The supernatant was removed leaving a whitish fluid containing clear cells that were used for “patch-clamp” recording of membrane ionic currents and potentials. Resting membrane potentials (RMPs) and action potentials of the cell membranes were measured using the whole-cell current mode. Glass microelectrodes, with a tip diameter 2–3 μm and tip resistance 5–6 MΩ (when filled with intracellular solution), were used. The mean values for RMP action potential and its duration showed no significant changes with the season, but other components of action potential, including threshold potential, after-potential, and its duration, were found to increase significantly (p < 0.05) in summer as compared to winter. This is the first study of seasonal temperature dependency of these parameters in the gastrocnemius muscle of U. hardwickii. This study shows that seasonal changes in components of the action potential are associated with changes in environmental temperature. These changes may be responsible for altered activities and homeostasis in these animals, and possibly related to the underlying mechanisms responsible for hibernation.

Using Higuchi’s fractal dimension in fine analysis of the effects of 2.7 mT and 10 mT static magnetic fields on the complex bursting activity of the snail BR neuron

The components of complex bioelectrical activity ? action potential (AP), interspike interval (ISI) and the quiet interburst interval (IBI), along with the effects of 2.7 mT and 10 mT static magnetic fields, were identified and examined in the spontaneously active Br neuron of the subesophageal ganglion complex of the garden snail Helix pomatia by fractal analysis using Higuchi?s fractal dimension (FD). The normalized mean of the empirical FD distributions of bursting activity of the Br neuron were formed under different experimental conditions: before (Control), during (MF), and after exposure to the static magnetic field (AMF). Using the fractal analysis method for the first time, a separation of the AP, ISI and IBI components was successfully achieved. Our results show that fractal analysis with deconvolution of the normalized mean of the empirical FD distributions into Gaussian functions is a useful tool for revealing the effects of magnetic fields on the complexity of neuronal bioelectric activity and its AP, ISI and IBI components.

Ouabain modulation of snail Br neuron bursting activity after the exposure to 10 mT static magnetic field revealed by Higuchi fractal dimension.

Aim of this study was to investigate the application of normalized mean of the empirical Higuchi fractal dimension (FD) distributions, as a new approach to analyze the spontaneous bioelectrical activity of garden snail (Helix pomatia) Br neuron. The effect of ouabain on modulation of Br neuron bursting activity before and after the exposure to 10 mT static magnetic field (SMF) was observed by analyzing the following parameters: action potential (AP), interspike interval (ISI) and interbursting interval (IBI) components. Normalized mean of the empirical FD distributions were formed for the following experimental conditions: Control 1, Ouabain 1, Control 2, SMF 2, ASMF 2, Control 3, SMF 3 and Ouabain ASMF 3. Our main results have shown that ouabain without SMF induced increase in participation of AP and a decrease in participation of IBI components compared to the first control condition. However, in the presence of 10 mT SMF, ouabain-induced changes of measured parameters of Br neuron activity were less pronounced compared to the third control condition. We have shown that normalized mean of the empirical FD distributions is a useful method for detecting the changes in AP, ISI, and IBI components of complex bursting activity in altered physiological states.

Reply from Dario Farina, Francesco Negro and Ning Jiang

Reply from Dario Farina, Francesco Negro and Ning Jiang

Decreased afferent excitability contributes to synaptic depression during high-frequency stimulation in hippocampal area CA1

Long-term potentiation (LTP) is often induced experimentally by continuous high-frequency afferent stimulation (HFS), typically at 100 Hz for 1 s. Induction of LTP requires postsynaptic depolarization and voltage-dependent calcium influx. Induction is more effective if the same number of stimuli are given as a series of short bursts rather than as continuous HFS, in part because excitatory postsynaptic potentials (EPSPs) become strongly depressed during HFS, reducing postsynaptic depolarization. In this study, we examined mechanisms of EPSP depression during HFS in area CA1 of rat hippocampal brain slices. We tested for presynaptic terminal vesicle depletion by examining minimal stimulation-evoked excitatory postsynaptic currents (EPSCs) during 100-Hz HFS. While transmission failures increased, consistent with vesicle depletion, EPSC latencies also increased during HFS, suggesting a decrease in afferent excitability. Extracellular recordings of Schaffer collateral fiber volleys confirmed a decrease in afferent excitability, with decreased fiber volley amplitudes and increased latencies during HFS. To determine the mechanism responsible for fiber volley changes, we recorded antidromic action potentials in single CA3 pyramidal neurons evoked by stimulating Schaffer collateral axons. During HFS, individual action potentials decreased in amplitude and increased in latency, and these changes were accompanied by a large increase in the probability of action potential failure. Time derivative and phase-plane analyses indicated decreases in both axon initial segment and somato-dendritic components of CA3 neuron action potentials. Our results indicate that decreased presynaptic axon excitability contributes to depression of excitatory synaptic transmission during HFS at synapses between Schaffer collaterals and CA1 pyramidal neurons.

Anatomic Localization and Autonomic Modulation of Atrioventricular Junctional Rhythm in Failing Human Hearts

The structure-function relationship in the atrioventricular junction (AVJ) of various animal species has been investigated in detail; however, less is known about the human AVJ. In this study, we performed high-resolution optical mapping of the human AVJ (n = 6) to define its pacemaker properties and response to autonomic stimulation. Isolated, coronary-perfused AVJ preparations from failing human hearts (n = 6, 53 ± 6 years) were optically mapped using the near-infrared, voltage-sensitive dye, di-4-ANBDQBS, with isoproterenol (1 μmol/L) and acetylcholine (1 μmol/L). An algorithm detecting multiple components of optical action potentials was used to reconstruct multilayered intramural AVJ activation and to identify specialized slow and fast conduction pathways (SP and FP). The anatomic origin and propagation of pacemaker activity was verified by histology. Spontaneous AVJ rhythms of 29 ± 11 bpm (n = 6) originated in the nodal-His region (n = 3) and/or the proximal His bundle (n = 4). Isoproterenol accelerated the AVJ rhythm to 69 ± 12 bpm (n = 5); shifted the leading pacemaker to the transitional cell regions near the FP and SP (n = 4) and/or coronary sinus (n = 2); and triggered reentrant arrhythmias (n = 2). Acetylcholine (n = 4) decreased the AVJ rhythm to 18 ± 4 bpm; slowed FP/SP conduction leading to block between the AVJ and atrium; and shifted the pacemaker to either the transitional cell region or the nodal-His region (bifocal activation). We have demonstrated that the AVJ pacemaker in failing human hearts is located in the nodal-His region or His bundle regions and can be modified with autonomic stimulation. Moreover, we found that both the FP and SP are involved in anterograde and retrograde conduction.

Effect of cold and menthol on membrane potential in plants

In animals, cooling substances such as menthol are perceived as cold sensation because they bind to the same receptor TRPM8 (transient receptor potential melastatin) that activates upon temperature drops. We investigated the effect of menthol on the plant membrane potential to search for analogies between animal and plant perception systems. The study was conducted on the liverwort Conocephalum conicum- a non-vascular plant generating action potentials (APs) in response to different stimuli including cold. (+)Menthol, (-)menthol and (+/-)menthol induced one or more APs, depending on the concentration. In contrast to animal reactions to menthol, threshold concentrations of these isomers were the same (1 mM). The presence of menthol in medium shortened cold-induced APs, whereas low temperature prolonged the repolarization phase of AP evoked by menthol. Cells of C. conicum with anion and potassium channels blocked by anthracene-9-carboxylic acid (A9C) and tetraethylammonium chloride (TEACl) generate short spike-like voltage transients (VTs) in response to cold and light stimulation. Membrane potential changes evoked by menthol in A9C- and TEACl-treated plants differed significantly from VTs - lasted much longer and frequently occurred in series. 5 mM LaCl(3) , 1 mM EGTA (ethylene glycol-bis(2-aminoethyl ether)-N,N,N',N'-tetraacetic acid) (0 Ca(2+) ) but not 0.2 mM verapamil blocked the putative calcium component of AP induced by menthol. Similar inhibitory effect was observed after the application of proton pump inhibitors: 0.05 mM N,N-dicyclohexylcarbodiimide (DCCD), 0.05 mM diethylstilbestrol (DES) or 0.01 mM carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP). Our results indicate that cold and menthol act independently, activating different membrane transporters in C. conicum cells.

Complex Interactions Between the Sinoatrial Node and Atrium During Reentrant Arrhythmias in the Canine Heart

Numerous studies implicate the sinoatrial node (SAN) as a participant in atrial arrhythmias, including atrial flutter (AFL) and atrial fibrillation (AF). However, the direct role of the SAN has never been described. The SAN was optically mapped in coronary perfused preparations from normal canine hearts (n=17). Optical action potentials were recorded during spontaneous rhythm, overdrive atrial pacing, and AF/AFL induced by acetylcholine (ACh; 0.3 to 3 micromol/L) and/or isoproterenol (Iso; 0.2 to 1 micromol/L). An optical action potential multiple component algorithm and dominant frequency analysis were used to reconstruct SAN activation and to identify specialized sinoatrial conduction pathways. Both ACh and Iso facilitated pacing-induced AF/AFL by shortening atrial repolarization. The entire SAN structure created a substrate for macroreentry with 9.6+/-1.7 Hz (69 episodes in all preparations). Atrial excitation waves could enter the SAN through the sinoatrial conduction pathways and overdrive suppress the node. The sinoatrial conduction pathways acted as a filter for atrial waves by slowing conduction and creating entrance block. ACh/Iso modulated filtering properties of the sinoatrial conduction pathways by increasing/decreasing the degree of the entrance block, respectively. Thus, the SAN could beat independently from AF/AFL reentrant activity during ACh (49+/-39%) and ACh/Iso (62+/-25%) (P=0.38). Without ACh, the AF/AFL waves captured the SAN and overdrive suppressed it. Spontaneous SAN activity could terminate or convert AFL to AF during cholinergic withdrawal. The specialized structure of the SAN can be a substrate for AF/AFL. Cholinergic stimulation not only can slow sinus rhythm and facilitate AF/AFL but also protects the intrinsic SAN function from the fast AF/AFL rhythm.

Linalool blocks excitability in peripheral nerves and voltage-dependent Na + current in dissociated dorsal root ganglia neurons

Linalool is a terpene that occurs as a major constituent of essential oils of many plants of widespread distribution. It possesses several biological and pharmacological activities, including depressant effects on the central nervous system and olfactory receptors. The present study investigated whether linalool affects the excitability of peripheral components of the somatic sensory system. We used sciatic nerve and preparations of intact and dissociated neurons of dorsal root ganglion for extracellular, intracellular and patch-clamp recordings. Linalool concentration-dependently (0.3–2.0 mM) and reversibly blocked the excitability of the sciatic nerve. It inhibited peak-to-peak amplitude of the compound action potential (IC 50 was 0.78 ± 0.04 mM). At 0.8 mM, it reversibly increased rheobase and chronaxy (from 3.2 ± 0.1 V and 52.4 ± 4.1 μs to 4.2 ± 0.3 V and 71.2 ± 5.5 μs (n = 5), respectively) and inhibited with greater pharmacological potency the amplitude of the compound action potential components corresponding to axons with slower velocity of conduction. In a similar concentration range (0.1–6 mM), linalool concentration-dependently and reversibly blocked the generation of action potentials of intact dorsal root ganglion neurons without alteration of resting membrane potential and input resistance, and inhibited the voltage-gated Na + current of dissociated dorsal root ganglion neurons. In conclusion, we demonstrated that linalool acts on the somatic sensory system with local anesthetic properties, since it blocked the action potential by acting on voltage-dependent Na + channels. This finding is important in showing the potential usefulness of linalool as a pharmacotherapeutic agent.

Electrophysiological and theoretical analysis of melatonin in peripheral nerve crush injury

Electrophysiological and theoretical studies have been performed to investigate peripheral nerve injuries and nerve regeneration. The aim of this work is to evaluate the effect of melatonin functionally and electrophysiologically on peripheral nerve crush injury. Adult male Wistar rats ( n = 32, 200 ± 50 g) were randomly allocated into four groups. Sciatic crush was constituted on left sciatic nerves. Treatment groups received intraperitoneal melatonin at doses of 5 and 20 mg/kg for 21 days. Functional nerve recovery was evaluated using sciatic functional index (SFI) every week during the experiment. In vivo electrophysiological measurements were performed at the end of the treatment. The electrophysiological data were also analyzed by wavelet analysis. Melatonin treatments increased the SFI values in the injured sciatic nerves. In vivo electrophysiological measurements showed that melatonin increased the conduction velocities and also decreased the latency values. The wavelet analysis showed that melatonin treatment reduced the densities of high frequency components of compound muscle action potential (CMAP). These results suggest that melatonin application is a promising strategy for the treatment of peripheral nerve crush injuries. Furthermore, analysis of EMG data with wavelet methods seems to give more reliable results to evaluate the nerve recovery.

Structural and Functional Evidence for Discrete Exit Pathways That Connect the Canine Sinoatrial Node and Atria

Surface electrode recordings cannot delineate the activation within the human or canine sinoatrial node (SAN) because they are intramural structures. Thus, the site of origin of excitation and conduction pathway(s) within the SAN of these mammals remains unknown. Canine right atrial preparations (n=7) were optically mapped. The SAN 3D structure and protein expression were mapped using immunohistochemistry. SAN optical action potentials had diastolic depolarization and multiple upstroke components that corresponded to the separate excitations of the node and surface atrial layers. Pacing-induced SAN exit block eliminated atrial optical action potential components but retained SAN optical action potential components. Excitation originated in the SAN (cycle length, 557+/-72 ms) and slowly spread (1.2 to 14 cm/sec) within the SAN, failing to directly excite the crista terminalis and intraatrial septum. After a 49+/-22 ms conduction delay within the SAN, excitation reached the atrial myocardium via superior and/or inferior sinoatrial exit pathways 8.8+/-3.2 mm from the leading pacemaker site. The ellipsoidal 13.7+/-2.8/4.9+/-0.6 mm SAN structure was functionally insulated from the atrium. This insulation coincided with connexin43-negative regions at the borders of the node, connective tissue, and coronary arteries. During normal sinus rhythm, the canine SAN is functionally insulated from the surrounding atrial myocardium except for 2 (or more) narrow superior and inferior sinoatrial exit pathways separated by 12.8+/-4.1 mm. Conduction failure in these sinoatrial exit pathways leads to SAN exit block and is a modulator of heart rate.

The Effects of pH on Beta-Endorphin and Morphine Inhibition of Calcium Transients in Dorsal Root Ganglion Neurons

During inflammation, immune cells migrate into inflamed tissue and release opioid peptides that activate opioid receptors on peripheral sensory neurons to reduce pain. A characteristic of the inflamed environment in which these opioids act is acidic pH. Activation of opioid receptors leads to a decrease in the calcium component of neuronal action potentials. We investigated the hypothesis that inhibitory effects of opioids on intracellular calcium transients in dorsal root ganglion neuronal cultures are potentiated at acidic extracellular pH. Intracellular calcium responses to stimulation with capsaicin were measured in untreated neurons or after preincubation with β-endorphin or morphine. β-Endorphin significantly inhibited calcium responses to 300 nmol/L capsaicin at the lowest experimental extracellular pH (6.1, 6.5, and 7.2), whereas morphine inhibited capsaicin (300 nmol/L) responses significantly at pH 6.1 with a trend of inhibition at pH 6.5. The effect of pH on morphine inhibition of K +-evoked calcium responses was also assessed. Morphine inhibition of calcium responses was significantly enhanced at pH 6.8 compared with pH 7.2 and pH 7.6. The inhibitory effects were reversed by naloxone, an opioid receptor antagonist. In conclusion, low extracellular pH potentiated β-endorphin and morphine inhibition of calcium transients and might contribute to improved opioid efficacy during inflammation. Perspective The results of the current study suggest that acidic pH might contribute to increased opioid efficacy in inflamed tissue. This highlights the therapeutic potential of endogenous opioid analgesia, whereby opioid peptides are delivered locally in inflamed tissues, as well as the use of locally applied opioids in inflammatory conditions.