scholarly journals Serotonergic Modulation of Persistent Inward Currents in Serotonergic Neurons of Medulla in ePet-EYFP Mice

2021 ◽  
Vol 15 ◽  
Author(s):  
Yi Cheng ◽  
Nan Song ◽  
Renkai Ge ◽  
Yue Dai
Keyword(s):  
Morphological Data ◽  
Repetitive Firing ◽  
Patch Clamp Technique ◽  
Serotonergic Neurons ◽  
Persistent Inward Currents ◽  
Functional Roles ◽  
Whole Cell Patch Clamp ◽  
Significant Difference ◽  
Inward Currents ◽  
Serotonergic Modulation

Serotonergic (5-HT) neurons in the medulla play multiple functional roles associated with many symptoms and motor activities. The descending serotonergic pathway from medulla is essential for initiating locomotion. However, the ionic properties of 5-HT neurons in the medulla remain unclear. Using whole-cell patch-clamp technique, we studied the biophysical and modulatory properties of persistent inward currents (PICs) in 5-HT neurons of medulla in ePet-EYFP transgenic mice (P3–P6). PICs were recorded by a family of voltage bi-ramps (10-s duration, 40-mV peak step), and the ascending and descending PICs were mirrored to analyze the PIC hysteresis. PICs were found in 77% of 5-HT neurons (198/258) with no significant difference between parapyramidal region (n = 107) and midline raphe nuclei (MRN) (n = 91) in either PIC onset (−47.4 ± 10 mV and −48.7 ± 7 mV; P = 0.44) or PIC amplitude (226.9 ± 138 pA and 259.2 ± 141 pA; P = 0.29). Ninety-six percentage (191/198) of the 5-HT neurons displayed counterclockwise hysteresis and four percentage (7/198) exhibited the clockwise hysteresis. The composite PICs could be differentiated as calcium component (Ca_PIC) by bath application of nimodipine (25 μM), sodium component (Na_PIC) by tetrodotoxin (TTX, 2 μM), and TTX- and dihydropyridine-resistance component (TDR_PIC) by TTX and nimodipine. Ca_PIC, Na_PIC and TDR_PIC all contributed to upregulation of excitability of 5-HT neurons. 5-HT (15 μM) enhanced the PICs, including a 26% increase in amplitude of the compound currents of Ca_PIC and TDR_PIC (P < 0.001, n = 9), 3.6 ± 5 mV hyperpolarization of Na_PIC and TDR_PIC onset (P < 0.05, n = 12), 30% increase in amplitude of TDR_PIC (P < 0.01), and 2.0 ± 3 mV hyperpolarization of TDR_PIC onset (P < 0.05, n = 18). 5-HT also facilitated repetitive firing of 5-HT neurons through modulation of composite PIC, Na_PIC and TDR_PIC, and Ca_PIC and TDR_PIC, respectively. In particular, the high voltage-activated TDR_PIC facilitated the repetitive firing in higher membrane potential, and this facilitation could be amplified by 5-HT. Morphological data analysis indicated that the dendrites of 5-HT neurons possessed dense spherical varicosities intensively crossing 5-HT neurons in medulla. We characterized the PICs in 5-HT neurons and unveiled the mechanism underlying upregulation of excitability of 5-HT neurons through serotonergic modulation of PICs. This study provided insight into channel mechanisms responsible for the serotonergic modulation of serotonergic neurons in brainstem.

Sequential activation of multiple persistent inward currents induces staircase currents in serotonergic neurons of medulla in ePet-EYFP mice

2020 ◽  
Vol 123 (1) ◽  
pp. 277-288
Author(s):  
Yi Cheng ◽  
Qiang Zhang ◽  
Yue Dai
Keyword(s):  
High Voltage ◽  
Low Voltage ◽  
Spinal Neurons ◽  
Serotonergic Neurons ◽  
Persistent Inward Currents ◽  
Functional Roles ◽  
Bath Application ◽  
Voltage Ramp ◽  
Inward Currents ◽  
Sequential Activation

Persistent inward currents (PICs) are widely reported in rodent spinal neurons. A distinctive pattern observed recently is staircase-like PICs induced by voltage ramp in serotonergic neurons of mouse medulla. The mechanism underlying this pattern of PICs is unclear. Combining electrophysiological, pharmacological, and computational approaches, we investigated the staircase PICs in serotonergic neurons of medulla in ePet-EYFP transgenic mice (postnatal days 1–7). Staircase PICs induced by 10-s voltage biramps were observed in 70% of serotonergic neurons ( n = 73). Staircase PICs activated at −48.8 ± 5 mV and consisted of two components, with the first PIC of 45.8 ± 51 pA and the second PIC of 197.3 ± 126 pA ( n = 51). Staircase PICs were also composed of low-voltage-activated sodium PIC (Na-PIC; onset −46.2 ± 5 mV, n = 34), high-voltage-activated calcium PIC (Ca-PIC; onset −29.3 ± 6 mV, n = 23), and high-voltage-activated tetrodotoxin (TTX)- and dihydropyridine-resistant sodium PIC (TDR-PIC; onset −16.8 ± 4 mV, n = 28). Serotonergic neurons expressing Na-PIC, Ca-PIC, and TDR-PIC were evenly distributed in medulla. Bath application of 1–2 μM TTX blocked the first PIC and decreased the second PIC by 36% ( n = 23, P < 0.05). Nimodipine (25 μM) reduced the second PIC by 38% ( n = 34, P < 0.001) without altering the first PIC. TTX and nimodipine removed the first PIC and reduced the second PIC by 59% ( n = 28, P < 0.01). A modeling study mimicked the staircase PICs and verified experimental conclusions that sequential activation of Na-PIC, Ca-PIC, and TDR-PIC in order of voltage thresholds induced staircase PICs in serotonergic neurons. Further experimental results suggested that the multiple components of staircase PICs play functional roles in regulating excitability of serotonergic neurons in medulla. NEW & NOTEWORTHY Staircase persistent inward currents (PICs) are mediated by activation of L-type calcium channels in dendrites of mouse spinal motoneurons. A novel mechanism is explored in this study. Here we report that the staircase PICs are mediated by sequentially activating sodium and calcium PICs in serotonergic neurons of mouse medulla.

Two distinct inactivation processes related to phosphorylation in cardiac L-type Ca2+ channel currents

2000 ◽  
Vol 279 (3) ◽  
pp. C603-C610 ◽  
Author(s):  
Sayaka Mitarai ◽  
Muneshige Kaibara ◽  
Katsusuke Yano ◽  
Kohtaro Taniyama
Keyword(s):  
Membrane Potential ◽  
Protein Kinase ◽  
Time Constant ◽  
Kinase Inhibitor ◽  
Slow Component ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Inward Currents ◽  
Inactivation Process ◽  
Channel Currents

We investigated the inactivation process of macroscopic cardiac L-type Ca2+ channel currents using the whole cell patch-clamp technique with Na+ as the current carrier. The inactivation process of the inward currents carried by Na+ through the channel consisted of two components >0 mV. The time constant of the faster inactivating component (30.6 ± 2.2 ms at 0 mV) decreased with depolarization, but the time constant of the slower inactivating component (489 ± 21 ms at 0 mV) was not significantly influenced by the membrane potential. The inactivation process in the presence of isoproterenol (100 nM) consisted of a single component (538 ± 60 ms at 0 mV). A protein kinase inhibitor, H-89, decreased the currents and attenuated the effects of isoproterenol. In the presence of cAMP (500 μM), the inactivation process consisted of a single slow component. We propose that the faster inactivating component represents a kinetic of the dephosphorylated or partially phosphorylated channel, and phosphorylation converts the kinetics into one with a different voltage dependency.

Expression of functional nicotinic acetylcholine receptors in neuroepithelial bodies of neonatal hamster lung

2003 ◽  
Vol 285 (6) ◽  
pp. L1203-L1212 ◽  
Author(s):  
Xiao Wen Fu ◽  
Colin A. Nurse ◽  
Suzanne M. Farragher ◽  
Ernest Cutz
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Respiratory Control ◽  
Hill Coefficient ◽  
Neuroepithelial Bodies ◽  
Patch Clamp Technique ◽  
Nicotinic Acetylcholine ◽  
Whole Cell Patch Clamp ◽  
Inward Currents ◽  
Double Label

Pulmonary neuroepithelial bodies (NEB) are presumed airway chemoreceptors involved in respiratory control, especially in the neonate. Nicotine is known to affect both lung development and control of breathing. We report expression of functional nicotinic acetylcholine receptors (nAChR) in NEB cells of neonatal hamster lung using a combination of morphological and electrophysiological techniques. Nonisotopic in situ hybridization method was used to localize mRNA for the β2-subunit of nAChR in NEB cells. Double-label immunofluorescence confirmed expression of α4-, α7-, and β2-subunits of nAChR in NEB cells. The electrophysiological characteristics of nAChR in NEB cells were studied using the whole cell patch-clamp technique on fresh lung slices. Application of nicotine (∼0.1-100 μM) evoked inward currents that were concentration dependent (EC50 = 3.8 μM; Hill coefficient = 1.1). ACh (100 μM) and nicotine (50 μM) produced two types of currents. In most NEB cells, nicotine-induced currents had a single desensitizing component that was blocked by mecamylamine (50 μM) and dihydro-β-erythroidine (50 μM). In some NEB cells, nicotine-induced current had two components, with fast- and slow-desensitizing kinetics. The fast component was selectively blocked by methyllcaconitine (MLA, 10 nM), whereas both components were inhibited by mecamylamine. Choline (0.5 mM) also induced an inward current that was abolished by 10 nM MLA. These studies suggest that NEB cells in neonatal hamster lung express functional heteromeric α3β2, α4β2, and α7 nAChR and that cholinergic mechanisms could modulate NEB chemoreceptor function under normal and pathological conditions.

Serotonergic modulation of neuronal activity in rat midbrain periaqueductal gray

2013 ◽  
Vol 109 (11) ◽  
pp. 2712-2719 ◽  
Author(s):  
Hyo-Jin Jeong ◽  
Karen Lam ◽  
Vanessa A. Mitchell ◽  
Christopher W. Vaughan
Keyword(s):  
Brain Slices ◽  
Periaqueductal Gray ◽  
Membrane Excitability ◽  
Inhibitory Effects ◽  
Whole Cell Patch Clamp ◽  
Outward Currents ◽  
Selective Ligands ◽  
Inward Currents ◽  
Subtype Selective ◽  
Serotonergic Modulation

Serotonin (5-HT) modulates pain and anxiety from within the midbrain periaqueductal gray (PAG). In the present study, the effects of 5-HT- and 5-HT1/2 subtype-selective ligands on rat PAG neurons were examined using whole cell patch-clamp recordings in brain slices. In voltage clamp, 5-HT produced outward and inward currents in distinct subpopulations of neurons that varied throughout different subregions of the PAG. The 5-HT1A agonist R(+)-8-OH-DPAT (1 μM) produced outward currents in subpopulations of PAG neurons. By contrast, sumatriptan (1 μM) and other 5-HT1B, -D, and -F subtype agonists had little or no postsynaptic activity. The 5-HT2A/C agonists DOI (3 μM) and TCB-2 (1 μM) produced inward currents in subpopulations of PAG neurons, and DOI enhanced evoked inhibitory postsynaptic currents via a presynaptic mechanism. In current clamp, both R(+)-8-OH-DPAT and sumatriptan produced an excitatory increase in evoked mixed postsynaptic potentials (PSPs). In addition, R(+)-8-OH-DPAT, but not sumatriptan, directly hyperpolarized PAG neurons. By contrast, the 5-HT2 agonist DOI depolarized subpopulations of neurons and produced an inhibitory decrease in evoked mixed PSPs. These findings indicate that 5-HT1A and 5-HT1B/D ligands have partly overlapping inhibitory effects on membrane excitability and synaptic transmission within the PAG, which are functionally opposed by 5-HT2A/C actions in specific PAG subregions.

Inhibition of glutamatergic transmission and neuronal excitability by oxycodone in the rat hippocampal CA3 neurons

2020 ◽  
Author(s):  
Cheng-Wei Lu ◽  
Tzu-Yu Lin ◽  
Shu-Kuei Huang ◽  
Kuan-Ming Chiu ◽  
Ming-Yi Lee ◽  
...  
Keyword(s):  
Neuronal Excitability ◽  
Glutamate Release ◽  
Opioid Analgesic ◽  
Inhibitory Effect ◽  
Repetitive Firing ◽  
Whole Cell Patch Clamp ◽  
Pain Transmission ◽  
Inward Currents ◽  
Hippocampal Ca3 ◽  
Ca3 Neurons

Oxycodone, a semisynthetic opioid analgesic with actions similar to morphine, is extensively prescribed for treatment of moderate to severe acute pain. Given that glutamate plays a crucial role in mediating pain transmission, the propose of this study was to investigate the effect of oxycodone on glutamatergic synaptic transmission in rat hippocampal CA3 area, which is associated with the modulation of nociceptive perception. Whole-cell patch-clamp recordings revealed that oxycodone effectively reduced presynaptic glutamate release, as detected by decreased frequencies of spontaneous excitatory postsynaptic currents (sEPSCs) and miniature EPSCs (mEPSCs), without eliciting significant changes in the amplitudes of sEPSCs and mEPSCs and glutamate-evoked inward currents. The inhibitory effect of oxycodone on the frequency of sEPSCs was blocked by the nonselective opioid receptor antagonist naloxone. In addition, oxycodone suppressed burst firing induced by 4-aminopyridine and tonic repetitive firing evoked by the applied depolarizing current. These results suggest that oxycodone inhibits spontaneous presynaptic glutamate release possibly by activating opioid receptors and consequently suppressing the neuronal excitability of hippocampal CA3 neurons.

scholarly journals Neuroepithelial bodies in mammalian lung express functional serotonin type 3 receptor

2001 ◽  
Vol 281 (4) ◽  
pp. L931-L940 ◽  
Author(s):  
X. W. Fu ◽  
D. Wang ◽  
J. Pan ◽  
S. M. Farragher ◽  
V. Wong ◽  
...  
Keyword(s):  
Functional Characterization ◽  
Dependent Manner ◽  
Induced Current ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Whole Cell Patch Clamp ◽  
Cell Plasma ◽  
Neuroepithelial Body ◽  
Inward Currents ◽  
Type 3

Serotonin (5-HT) type 3 receptor (5-HT3-R) is a ligand-gated ion channel found primarily in the central and peripheral nervous system. We report expression and functional characterization of 5-HT3-R in pulmonary neuroepithelial body (NEB) cells. Using nonisotopic in situ hybridization, we demonstrate expression of 5-HT3-R mRNA in NEB cells in the lungs of different mammals (hamster, rabbit, mouse, and human). Dual immunocytochemistry (for 5-HT and 5-HT3-R) and confocal microscopy localized 5-HT3-R on NEB cell plasma membrane from rabbit. The electrophysiological characteristics of 5-HT3-R in NEB cells were studied in fresh slices of neonatal hamster lung using the whole cell patch-clamp technique. Application of the 5-HT (5–150 μM) and 5-HT3-R agonist 2-methyl-5-HT (5–150 μM) induced inward currents in a concentration-dependent manner. The 5-HT-induced current was blocked (76.5 ± 5.9%) by the specific 5-HT3-R antagonist ICS-205–930 (50 μM), whereas katanserin and p-4-iodo- N-{2-[4-(methoxyphenyl)-1-piperazinyl]ethyl}- N-2-pyridinylbenzamide had minimal effects. Forskolin had no effect on desensitization and amplitude of the 5-HT-induced current. The reduction of Ca2+ and Mg2+ in the extracellular solution enhanced the amplitude of the 5-HT-induced current because of slower desensitization. Our studies suggest that 5-HT3-R in NEB cells may function as an autoreceptor and may potentially be involved in modulation of hypoxia signaling.

GABA-Mimetic Actions of Withania somnifera on Substantia Gelatinosa Neurons of the Trigeminal Subnucleus Caudalis in Mice

2013 ◽  
Vol 41 (05) ◽  
pp. 1043-1051 ◽  
Author(s):  
Hua Yin ◽  
Dong Hyu Cho ◽  
Soo Joung Park ◽  
Seong Kyu Han
Keyword(s):  
Patch Clamp ◽  
Receptor Antagonist ◽  
Withania Somnifera ◽  
Substantia Gelatinosa ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
Inward Currents

The plant Withania somnifera (WS), also known as Ashwagandha, has been used widely in traditional medicine systems in India and Nepal (Ayurveda), and has been accepted to cure various ailments. In this study, the whole-cell patch clamp technique was performed to examine the mechanism of action of WS on the SG neurons of the Vc from mouse brainstem slices. In whole-cell patch clamp mode, methanol extract of Withania somnifera (mWS) induced short-lived and repeatable inward currents in all SG neurons tested (31.3±8.51 pA, n = 7) using a high chloride pipette solution. The mWS-induced inward currents were concentration dependent and maintained in the presence of tetrodotoxin (TTX), a voltage gated Na + channel blocker, CNQX, a non-NMDA glutamate receptor antagonist, AP5, an NMDA receptor antagonist and strychnine, a glycine receptor antagonist. The mWS induced currents were blocked by picrotoxin, a GABAA receptor antagonist. These results show that mWS has an inhibitory effects on SG neurons of the Vc through GABAA receptor-mediated activation of chloride ion channels, indicating that mWS contains compounds with sedative effects on the central nervous system. These results also suggest that mWS may be a potential target for modulating orofacial pain processing.

An unusual voltage-gated anion channel found in the cone cells of the chicken retina

1991 ◽  
Vol 6 (1) ◽  
pp. 19-23 ◽  
Author(s):  
Martin Wilson ◽  
Evanna Gleason
Keyword(s):  
Patch Clamp ◽  
Anion Channel ◽  
Chloride Ions ◽  
Patch Clamp Technique ◽  
Adult Chicken ◽  
Voltage Gated ◽  
Cationic Current ◽  
Whole Cell Patch Clamp ◽  
Cone Cells ◽  
Inward Currents

AbstractUsing the whole-cell patch clamp technique, we have examined the voltage-gated currents present in adult chicken cone cells. When calcium and calcium-gated currents are blocked, hyperpolarizing voltage steps elicit slowly increasing inward currents as has been shown for photoreceptors in other species. Unlike the case for other species, chicken cones appear to lack the inward-rectifying cationic current Ih that contributes to the shaping of the photovoltage. Instead of Ih, these cones possess an anionic inward-rectifying current that in kinetics, activation range and probably function is remarkably similar to Ih. This anion channel is unusual in that both nitrate and acetate are more permeant than chloride ions.

scholarly journals Maturation of persistent and hyperpolarization-activated inward currents shapes the differential activation of motoneuron subtypes during postnatal development

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Simon A Sharples ◽  
Gareth B Miles
Keyword(s):  
Postnatal Development ◽  
Motor Units ◽  
Intrinsic Properties ◽  
Size Principle ◽  
Differential Activation ◽  
Persistent Inward Currents ◽  
Poor Predictor ◽  
Whole Cell Patch Clamp ◽  
Inward Currents ◽  
Patch Clamp Electrophysiology

The size principle underlies the orderly recruitment of motor units; however, motoneuron size is a poor predictor of recruitment amongst functionally defined motoneuron subtypes. Whilst intrinsic properties are key regulators of motoneuron recruitment, the underlying currents involved are not well defined. Whole-cell patch-clamp electrophysiology was deployed to study intrinsic properties, and the underlying currents, that contribute to the differential activation of delayed and immediate firing motoneuron subtypes. Motoneurons were studied during the first three postnatal weeks in mice to identify key properties that contribute to rheobase and may be important to establish orderly recruitment. We find that delayed and immediate firing motoneurons are functionally homogeneous during the first postnatal week and are activated based on size, irrespective of subtype. The rheobase of motoneuron subtypes become staggered during the second postnatal week, which coincides with the differential maturation of passive and active properties, particularly persistent inward currents. Rheobase of delayed firing motoneurons increases further in the third postnatal week due to the development of a prominent resting hyperpolarization-activated inward current. Our results suggest that motoneuron recruitment is multifactorial, with recruitment order established during postnatal development through the differential maturation of passive properties and sequential integration of persistent and hyperpolarization-activated inward currents.

Human embryonic stem cell-derived cardiomyocytes: demonstration of a portion of cardiac cells with fairly mature electrical phenotype

2010 ◽  
Vol 235 (4) ◽  
pp. 522-530 ◽  
Author(s):  
Mari Pekkanen-Mattila ◽  
Hugh Chapman ◽  
Erja Kerkelä ◽  
Riitta Suuronen ◽  
Heli Skottman ◽  
...  
Keyword(s):  
Electrical Properties ◽  
Cell Model ◽  
Embryonic Stem ◽  
Cardiac Cells ◽  
Patch Clamp Technique ◽  
Drug Induced ◽  
Electrophysiological Properties ◽  
Promising Tool ◽  
Whole Cell Patch Clamp ◽  
Significant Difference

Cardiomyocytes (CMs) derived from human embryonic stem cells (hESC) provide a promising tool for the pharmaceutical industry. In this study the electrical properties and maturation of hESC-CM derived using two differentiation methods were compared and the suitability of hESC-CMs as a cell model for the assessment of drug-induced repolarization delay was evaluated. CMs were differentiated either in END-2 co-culture or by spontaneous differentiation. Action potentials (APs) were recorded from cells in spontaneously beating areas using the whole-cell patch-clamp technique. The hESC-CMs exhibited predominantly a ventricular-like phenotype with heterogeneous properties. Heterogeneity was indicative of the spectrum of hESC-CM maturation from embryonic-like with AP upstroke velocities <30 V/s and maximum diastolic potential (MDP) of close to −60 mV to more mature with values >150 V/s and −80 mV, respectively. The mean MDP was −70 mV and a significant difference was observed between the two differentiation methods (−66 versus −75 mV, P < 0.001). The age of the CMs did not correlate with phenotype maturation. The addition of the hERG blocker E-4031 and the sodium channel modulator veratridine significantly prolonged the AP duration. Furthermore, proarrhythmic indices were induced. In conclusion, the main observation was the heterogeneity in electrical properties of the hESC-CMs and this was observed with both differentiation methods. One-third of the hESC-CMs exhibited fairly mature electrophysiological properties, suggesting that mature CMs could be obtained from hESCs. However, improved differentiation methods are needed to produce homogeneous mature human CMs for pharmaceutical and toxicological applications.

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