026 Vasoactive Intestinal Polypeptide Directly Excites Neurons of the Subparaventricular Zone

SLEEP ◽  
2021 ◽  
Vol 44 (Supplement_2) ◽  
pp. A12-A12
Author(s):  
Francesca Raffin ◽  
Michela Cristofolini ◽  
Gerardo Rosario Biella ◽  
Patrick Fuller ◽  
Elda Arrigoni
Keyword(s):  
Patch Clamp ◽  
Vasoactive Intestinal Polypeptide ◽  
Calcium Imaging ◽  
Brain Slices ◽  
Membrane Depolarization ◽  
Patch Clamp Recording ◽  
Whole Cell ◽  
Subparaventricular Zone ◽  
Intestinal Polypeptide

Abstract Introduction The suprachiasmatic nucleus (SCN) is responsible for generating the circadian rhythmicity in mammals. The ventral region or core of the SCN contains neurons that express the neuropeptide vasoactive intestinal polypeptide (VIP). VIP signaling is central for coherency and synchrony of SCN activity. VIP-expressing neurons in the SCN densely project to the ventral subregions of the subparaventricular zone (vSPZ). We studied the effects of VIP on vSPZ neurons in brain slices of mice with a combined calcium imaging and whole-cell patch-clamp recording techniques. We used calcium imaging to assess the effects of VIP on vSPZ neurons as a population and we acquired patch-clamp recordings to explore the effects of VIP on the electrical properties and the synaptic inputs to vSPZ neurons. Methods We expressed GCamp6 in vSPZ neurons by stereotaxically injecting AAV10-DIO-Ef1a-GCamp6 into the vSPZ of vGAT-IRES-Cre mice. Brain slices were prepared two weeks later and images were captured using a standard GFP filter set. We performed whole-cell recordings of the vSPZ neurons of wild-type mice. We assessed the effects of VIP on the membrane potential and the on excitatory synaptic input in vSPZ neurons. Results Using GCamp6-based in vitro calcium imaging we found that VIP excites 17% of vSPZ neurons and this effect was maintained in the presence of tetrodotoxin (TTX) and synaptic blockers for AMPA/NMDA and GABAA transmissions suggesting a direct effect of VIP on vSPZ neurons. We confirmed this result with patch-clamp recordings. We found that 29% of vSPZ neurons were excited by VIP. VIP produced a membrane depolarization of vSPZ neurons in the presence of antagonists for AMPA/NMDA and GABAA receptors. In addition, we found that in a small percentage of vSPZ neurons VIP increased the frequency of the glutamatergic excitatory postsynaptic currents, suggesting an additional excitatory mechanism. Conclusion Our results demonstrate that exogenous VIP directly excites the vSPZ neurons producing an increase in intracellular calcium and membrane depolarization. In addition, VIP increases glutamatergic afferent inputs to vSPZ neurons indicating an additional synergistic excitation. We conclude that when VIP is released from the SCN VIP fibers it can activate vSPZ neurons. Support (if any) NS091126 and HL149630.

scholarly journals Local circuit input to the medullary reticular formation from the rostral nucleus of the solitary tract

2008 ◽  
Vol 295 (5) ◽  
pp. R1391-R1408 ◽  
Author(s):  
J. Nasse ◽  
D. Terman ◽  
S. Venugopal ◽  
G. Hermann ◽  
R. Rogers ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Reticular Formation ◽  
Calcium Imaging ◽  
Neonatal Rat ◽  
Solitary Tract ◽  
Patch Clamp Recording ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
Stimulation Of

The intermediate reticular formation (IRt) subjacent to the rostral (gustatory) nucleus of the solitary tract (rNST) receives projections from the rNST and appears essential to the expression of taste-elicited ingestion and rejection responses. We used whole cell patch-clamp recording and calcium imaging to characterize responses from an identified population of prehypoglossal neurons in the IRt to electrical stimulation of the rNST in a neonatal rat pup slice preparation. The calcium imaging studies indicated that IRt neurons could be activated by rNST stimulation and that many neurons were under tonic inhibition. Whole cell patch-clamp recording revealed mono- and polysynaptic projections from the rNST to identified prehypoglossal neurons. The projection was primarily excitatory and glutamatergic; however, there were some inhibitory GABAergic projections, and many neurons received excitatory and inhibitory inputs. There was also evidence of disinhibition. Overall, bath application of GABAA antagonists increased the amplitude of excitatory currents, and, in several neurons, stimulation of the rNST systematically decreased inhibitory currents. We have hypothesized that the transition from licks to gapes by natural stimuli, such as quinine monohydrochloride, could occur via such disinhibition. We present an updated dynamic model that summarizes the complex synaptic interface between the rNST and the IRt and demonstrates how inhibition could contribute to the transition from ingestion to rejection.

scholarly journals Automatic deep learning driven label-free image guided patch clamp system for human and rodent in vitro slice physiology

2020 ◽  
Author(s):  
Krisztian Koos ◽  
Gáspár Oláh ◽  
Tamas Balassa ◽  
Norbert Mihut ◽  
Márton Rózsa ◽  
...  
Keyword(s):  
Image Analysis ◽  
Deep Learning ◽  
Patch Clamp ◽  
Brain Research ◽  
Brain Slices ◽  
Cell Detection ◽  
Label Free ◽  
Patch Clamp Recording ◽  
Tissue Slices

ABSTRACTPatch clamp recording of neurons is a labor-intensive and time-consuming procedure. We have developed a tool that fully automatically performs electrophysiological recordings in label-free tissue slices. The automation covers the detection of cells in label-free images, calibration of the micropipette movement, approach to the cell with the pipette, formation of the whole-cell configuration, and recording. The cell detection is based on deep learning. The model was trained on a new image database of neurons in unlabeled brain tissue slices. The pipette tip detection and approaching phase use image analysis techniques for precise movements. High-quality measurements were performed on hundreds of human and rodent neurons. We also demonstrate that further molecular and anatomical analysis can be performed on the recorded cells. The software has a diary module that automatically logs patch clamp events. Our tool can multiply the number of daily measurements to help brain research.ONE SENTENCE SUMMARYNovel deep learning and image analysis algorithms for automated patch clamp systems to reliably measure neurons in human and rodent brain slices.

scholarly journals Disrupted Neuronal Activity Rhythms in the Suprachiasmatic Nuclei of Vasoactive Intestinal Polypeptide-Deficient Mice

2007 ◽  
Vol 97 (3) ◽  
pp. 2553-2558 ◽  
Author(s):  
T. M. Brown ◽  
C. S. Colwell ◽  
J. A. Waschek ◽  
H. D. Piggins
Keyword(s):  
Vasoactive Intestinal Polypeptide ◽  
Wheel Running ◽  
Brain Slices ◽  
Suprachiasmatic Nuclei ◽  
Wild Type ◽  
Behavioral Rhythms ◽  
Adult Mice ◽  
Deficient Mice ◽  
Intestinal Polypeptide

Vasoactive intestinal polypeptide (VIP), acting via the VPAC2 receptor, is a key signaling pathway in the suprachiasmatic nuclei (SCN), the master clock controlling daily rhythms in mammals. Most mice lacking functional VPAC2 receptors are unable to sustain behavioral rhythms and lack detectable SCN electrical rhythms in vitro. Adult mice that do not produce VIP (VIP/PHI−/−) exhibit less severe alterations in wheel-running rhythms, but the effects of this deficiency on the amplitude, phasing, or periodicity of their SCN cellular rhythms are unknown. To investigate this, we used suction electrodes to extracellularly record multiple- and single-unit electrical activity in SCN brain slices from mice with varying degrees of VIP deficiency, ranging from wild-type (VIP/PHI+/+) to heterozygous (VIP/PHI+/−) and VIP/PHI−/− animals. We found decreasing proportions of rhythmic cells in SCN slices from VIP/PHI+/+ (∼91%, n = 23) through VIP/PHI-/+ (∼71%, n = 28) to VIP/PHI−/− mice (62%; n = 37) and a parallel trend toward decreasing amplitude in the remaining rhythmic cells. SCN neurons from VIP/PHI−/− mice exhibited a broad range in the period and phasing of electrical rhythms, concordant with the known alterations in their behavioral rhythms. Further, treatment of VIP/PHI−/− slices with a VPAC2 receptor antagonist significantly reduced the proportion of oscillating neurons, suggesting that VPAC2 receptors still become activated in the SCN of these mice. The results establish that VIP is important for appropriate periodicity and phasing of SCN neuronal rhythms and suggest that residual VPAC2 receptor signaling promotes rhythmicity in adult VIP/PHI−/− mice.

scholarly journals Investigation of Neuron Latency Modulated by Bilateral Inferior Collicular Interactions Using Whole-Cell Patch Clamp Recording in Brain Slices

2021 ◽  
Vol 2021 ◽  
pp. 1-13
Author(s):  
Jinzhe Ma ◽  
Yangyang Han ◽  
Yiting Yao ◽  
Huimei Wang ◽  
Mengxia Chen ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Synaptic Input ◽  
Brain Slices ◽  
Inhibitory Neurons ◽  
Patch Clamp Recording ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
Inferior Collicular ◽  
Latency Variation

As the final level of the binaural integration center in the subcortical nucleus, the inferior colliculus (IC) plays an essential role in receiving binaural information input. Previous studies have focused on how interactions between the bilateral IC affect the firing rate of IC neurons. However, little is known concerning how the interactions within the bilateral IC affect neuron latency. In this study, we explored the synaptic mechanism of the effect of bilateral IC interactions on the latency of IC neurons. We used whole-cell patch clamp recordings to assess synaptic responses in isolated brain slices of Kunming mice. The results demonstrated that the excitation-inhibition projection was the main projection between the bilateral IC. Also, the bilateral IC interactions could change the reaction latency of most neurons to different degrees. The variation in latency was related to the type of synaptic input and the relative intensity of the excitation and inhibition. Furthermore, the latency variation also was caused by the duration change of the first subthreshold depolarization firing response of the neurons. The distribution characteristics of the different types of synaptic input also differed. Excitatory-inhibitory neurons were widely distributed in the IC dorsal and central nuclei, while excitatory neurons were relatively concentrated in these two nuclei. Inhibitory neurons did not exhibit any apparent distribution trend due to the small number of assessed neurons. These results provided an experimental reference to reveal the modulatory functions of bilateral IC projections.

Vasoactive intestinal polypeptide modulates GABAA receptor function in bipolar cells and ganglion cells of the rat retina

1992 ◽  
Vol 67 (4) ◽  
pp. 791-797 ◽  
Author(s):  
M. L. Veruki ◽  
H. H. Yeh
Keyword(s):  
Gabaa Receptor ◽  
Vasoactive Intestinal Polypeptide ◽  
Ganglion Cells ◽  
Bipolar Cells ◽  
Current Response ◽  
Gamma Aminobutyric Acid ◽  
Patch Clamp Recording ◽  
Rat Retina ◽  
Neuronal Populations ◽  
Intestinal Polypeptide

1. The effect of vasoactive intestinal polypeptide (VIP) on bipolar cells and ganglion cells freshly dissociated from the rat retina was studied under voltage clamp with the use of patch-clamp recording in the whole-cell configuration. 2. Application of VIP (1-100 microM) by itself resulted in no detectable current response in either bipolar cells or ganglion cells. However, gamma-aminobutyric acid (GABA)-activated macroscopic current responses elicited in both neuronal populations were potentiated on superimposed exposure to the neuropeptide. 3. GABA-activated chloride currents and muscimol-induced current responses were similarly potentiated on exposure to VIP, suggesting a synergistic interaction between VIP and GABAA receptor mechanisms. 4. We postulate that VIP plays a neuromodulatory role by regulating the excitability of inner retinal neurons and in this way modulates the efficacy of synaptic transmission in the retina.

scholarly journals Cholinergic Modulation of the Resonance Properties of Stellate Cells in Layer II of Medial Entorhinal Cortex

2010 ◽  
Vol 104 (1) ◽  
pp. 258-270 ◽  
Author(s):  
James G. Heys ◽  
Lisa M. Giocomo ◽  
Michael E. Hasselmo
Keyword(s):  
Patch Clamp ◽  
Resonance Frequency ◽  
Entorhinal Cortex ◽  
Stellate Cells ◽  
Cholinergic Modulation ◽  
Medial Entorhinal Cortex ◽  
Whole Cell ◽  
Whole Cell Patch Clamp ◽  
Resonance Properties

In vitro whole cell patch-clamp recordings of stellate cells in layer II of medial entorhinal cortex show a subthreshold membrane potential resonance in response to a sinusoidal current injection of varying frequency. Physiological recordings from awake behaving animals show that neurons in layer II medial entorhinal cortex, termed “grid cells,” fire in a spatially selective manner such that each cell's multiple firing fields form a hexagonal grid. Both the spatial periodicity of the grid fields and the resonance frequency change systematically in neurons along the dorsal to ventral axis of medial entorhinal cortex. Previous work has also shown that grid field spacing and acetylcholine levels change as a function of the novelty to a particular environment. Using in vitro whole cell patch-clamp recordings, our study shows that both resonance frequency and resonance strength vary as a function of cholinergic modulation. Furthermore, our data suggest that these changes in resonance properties are mediated through modulation of h-current and m-current.

Dopamine enhancement and depression of glutamate-regulated calcium and electrical activity in hypothalamic neurons

1996 ◽  
Vol 76 (6) ◽  
pp. 3934-3948 ◽  
Author(s):  
A. N. van den Pol ◽  
V. Cao ◽  
A. B. Belousov
Keyword(s):  
Membrane Potential ◽  
Electrical Activity ◽  
Dopamine Receptor ◽  
Glutamate Receptor ◽  
Independent Effect ◽  
Patch Clamp Recording ◽  
Hypothalamic Neurons ◽  
Whole Cell ◽  
Synaptic Release

1. The neurotransmitter dopamine is found throughout the hypothalamus both in cell bodies and in axons originating from intra- and extrahypothalamic sources. To study the mechanisms of action of dopamine on cultured rat hypothalamic neurons, particularly in relation to Ca2+ regulation, we used Ca2+ digital imaging with fura-2 and whole cell patch-clamp recording. We focused on the modulatory actions of dopamine on glutamate. 2. Dopamine administration had little or no independent effect on intracellular Ca2+. However, in the presence of tetrodotoxin to block action potentials and action-potential-dependent transmitter release, dopamine (10 microM for 2-3 min) caused an increase in glutamate-evoked Ca2+ rises in 22% of 64 neurons and depressed glutamate-evoked Ca2+ rises in an equal number of neurons. Shorter exposure to dopamine reduced the number of responding cells. 3. Dopamine application to neurons with an elevated Ca2+ due to synaptic release of glutamate (in the absence of tetrodotoxin) generally caused a decrease in Ca2+ levels (40% of 106 neurons), but sometimes increased cytosolic Ca2+ (10% of 106 neurons). That dopamine influenced cells differently in conditions of spontaneous activity compared with evoked activity may be due to dopamine effects on presynaptic receptors detected under conditions of ongoing synaptic release of glutamate. 4. Dopamine modulation of glutamate responses was detected at early stages of neuronal development (embryonic day 18 after 2 days in vitro) and also after 60 days in vitro. 5. The D1, D2, and D3 dopamine receptor agonists SKF38393, quinpirole, and 7-OH-DPAT (+/- 7 hydroxy-dipropylaminotetralin) caused a reduction in Ca2+ levels raised by endogenous glutamate release or evoked by exogenous glutamate application. 6. To block the actions of dopamine released by hypothalamic neurons, D1 and D2 dopamine receptor antagonists were used. As with dopamine, dopamine antagonists had no effect on intracellular Ca2+ during glutamate receptor blockade. In the absence of glutamate receptor block, the D1 antagonist SCH23390 (1 microM) reduced Ca2+ in responding cells; in contrast, the D2 antagonist eticlopride (1 microM) generated a delayed increase in Ca2+ levels. 7. Dopamine is known to activate second messengers through G proteins independent of changes in membrane potential or input resistance. Whole cell recording was used to demonstrate that, parallel to the modulation of Ca2+, dopamine exerted a dramatic change in glutamate-mediated electrical activity, generally depressing activity and hyperpolarizing the membrane potential (8 of 15 neurons). In a smaller number of neurons (5 of 15), dopamine enhanced glutamate-mediated excitatory activity. 8. Dopamine-evoked changes in membrane potential were in part mediated through modulation of glutamate actions. Dopamine depressed glutamate-evoked currents in a dose-dependent fashion, with Hill slopes in individual neurons ranging from 0.3 to 0.6. Dopamine could also evoke a direct hyperpolarizing action on hypothalamic neurons in the presence of tetrodotoxin or glutamate receptor blockers, at least in part by opening K+ channels. 9. Glutamate plays an important role as a primary excitatory transmitter within the hypothalamus. Our data support the hypothesis that a major mechanism of dopamine's influence on hypothalamic neurons involves the modulation of glutamate's excitatory action, mostly by inhibition. This is consistent with the hypothesis that modulation of glutamate activity may be an important mechanism of dopamine action throughout the nervous system.

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