scholarly journals Examinations of Bilateral Epileptiform Activities in Hippocampal Slices Obtained From Young Mice

2021 ◽  
Vol 14 ◽  
Author(s):  
Haiyu Liu ◽  
Peter L. Carlen ◽  
Liang Zhang
Keyword(s):  
Room Temperature ◽  
Hippocampal Slices ◽  
Field Potentials ◽  
Hippocampal Commissure ◽  
Extracellular Recordings ◽  
Adult Mice ◽  
Unilateral Stimulation ◽  
Dorsal Hippocampal ◽  
Young Mice

Bilateral interconnections through the hippocampal commissure play important roles in synchronizing or spreading hippocampal seizure activities. Intact hippocampi or bilateral hippocampal slices have been isolated from neonatal or immature rats (6–7 or 12–21 days old, respectively) and the mechanisms underlying the bilateral synchrony of hippocampal epileptiform activities have been investigated. However, the feasibility of examining bilateral epileptiform activities of more developed hippocampal circuitry in vitro remains to be explored. For this, we prepared bilateral hippocampal slices from C57 black mice, a strain commonly used in neuroscience and for genetic/molecular modifications. Young mice (21–24-day-old) were used in most experiments. A 600-μm-thick slice was obtained from each mouse by horizontal vibratome sectioning. Bilateral dorsal hippocampal and connecting dorsal hippocampal commissure (DHC) tissues were preserved in the slice and extrahippocampal tissues were removed. Slices were recorded in a submerged chamber mainly at a room temperature (21–22°C). Bilateral CA3 areas were monitored by extracellular recordings, and unilateral electrical stimulation was used to elicit CA3 synaptic field potentials. The unilateral stimulation could elicit population spikes in the contralateral CA3 area. These contralateral spikes were attenuated by inhibiting synaptic transmission with cobalt-containing medium and were abolished when a cut was made at the DHC. Self-sustained and bilaterally correlated epileptiform potentials were observed following application of 4-aminopyradine and became independent after the DHC cut. Bilateral hippocampal activities were detectable in some slices of adult mice and/or at 35–36°C, but with smaller amplitudes and variable waveforms compared to those observed from slices of young mice and at the room temperature. Together, these observations suggested that examining bilateral epileptiform activities in hippocampal slices of young mice is feasible. The weaknesses and limitations of this preparation and our experimentation are discussed.

scholarly journals Imaging calcium microdomains within entire astrocyte territories and endfeet with GCaMPs expressed using adeno-associated viruses

2013 ◽  
Vol 141 (5) ◽  
pp. 633-647 ◽  
Author(s):  
Eiji Shigetomi ◽  
Eric A. Bushong ◽  
Martin D. Haustein ◽  
Xiaoping Tong ◽  
Olan Jackson-Weaver ◽  
...  
Keyword(s):  
Blood Vessels ◽  
Hippocampal Slices ◽  
Adult Mice ◽  
Neuronal Synapses ◽  
Primary Signal ◽  
Experimental Approaches ◽  
Acute Hippocampal Slices

Intracellular Ca2+ transients are considered a primary signal by which astrocytes interact with neurons and blood vessels. With existing commonly used methods, Ca2+ has been studied only within astrocyte somata and thick branches, leaving the distal fine branchlets and endfeet that are most proximate to neuronal synapses and blood vessels largely unexplored. Here, using cytosolic and membrane-tethered forms of genetically encoded Ca2+ indicators (GECIs; cyto-GCaMP3 and Lck-GCaMP3), we report well-characterized approaches that overcome these limitations. We used in vivo microinjections of adeno-associated viruses to express GECIs in astrocytes and studied Ca2+ signals in acute hippocampal slices in vitro from adult mice (aged ∼P80) two weeks after infection. Our data reveal a sparkling panorama of unexpectedly numerous, frequent, equivalently scaled, and highly localized Ca2+ microdomains within entire astrocyte territories in situ within acute hippocampal slices, consistent with the distribution of perisynaptic branchlets described using electron microscopy. Signals from endfeet were revealed with particular clarity. The tools and experimental approaches we describe in detail allow for the systematic study of Ca2+ signals within entire astrocytes, including within fine perisynaptic branchlets and vessel-associated endfeet, permitting rigorous evaluation of how astrocytes contribute to brain function.

scholarly journals An In Vitro Model of Hippocampal Sharp Waves: Regional Initiation and Intracellular Correlates

2005 ◽  
Vol 94 (1) ◽  
pp. 741-753 ◽  
Author(s):  
Chiping Wu ◽  
Marjan Nassiri Asl ◽  
Jesse Gillis ◽  
Frances K. Skinner ◽  
Liang Zhang
Keyword(s):  
Dentate Gyrus ◽  
Large Amplitude ◽  
Pyramidal Neurons ◽  
Slow Wave Sleep ◽  
Hippocampal Slices ◽  
Field Potentials ◽  
Sharp Wave ◽  
Sharp Waves ◽  
Hippocampal Circuitry

During slow wave sleep and consummatory behaviors, electroencephalographic recordings from the rodent hippocampus reveal large amplitude potentials called sharp waves. The sharp waves originate from the CA3 circuitry and their generation is correlated with coherent discharges of CA3 pyramidal neurons and dependent on activities mediated by AMPA glutamate receptors. To model sharp waves in a relatively large hippocampal circuitry in vitro, we developed thick (1 mm) mouse hippocampal slices by separating the dentate gyrus from the CA2/CA1 areas while keeping the functional dentate gyrus-CA3-CA1 connections. We found that large amplitude (0.3–3 mV) sharp wave-like field potentials occurred spontaneously in the thick slices without extra ionic or pharmacological manipulation and they resemble closely electroencephalographic sharp waves with respect to waveform, regional initiation, pharmacological manipulations, and intracellular correlates. Through measuring tissue O2, K+, and synaptic and single cell activities, we verified that the sharp wave-like potentials are not a consequence of anoxia, nonspecific elevation of extracellular K+ and dissection-related tissue damage. Our data suggest that a subtle but crucial increase in the CA3 glutamatergic activity effectively recruits a population of neurons thus responsible for the generation of the sharp wave-like spontaneous field potentials in isolated hippocampal circuitry.

scholarly journals Stringent structural plasticity of dendritic spines revealed by two-photon glutamate uncaging in adult mouse neocortex in vivo

2019 ◽  
Author(s):  
Jun Noguchi ◽  
Akira Nagaoka ◽  
Tatsuya Hayama ◽  
Hasan Ucar ◽  
Sho Yagishita ◽  
...  
Keyword(s):  
Time Course ◽  
Hippocampal Slices ◽  
Low Frequency ◽  
Structural Plasticity ◽  
Two Photon ◽  
Low Magnesium ◽  
Adult Mice ◽  
First Time

AbstractTwo-photon uncaging of glutamate is widely utilized to characterize structural plasticity in brain slice preparations in vitro. In this study, we investigated spine plasticity by using, for the first time, glutamate uncaging in the neocortex of adult mice in vivo. Spine enlargement was successfully induced in a smaller fraction of spines in the neocortex (22%) than in young hippocampal slices (95%), even under a low magnesium condition. Once induced, the time course and mean amplitudes of long-term enlargement were the same (81%) as those in vitro. However, low-frequency (1–2 Hz) glutamate uncaging caused spine shrinkage in a similar fraction (34%) of spines as in vitro, but spread to the neighboring spines less frequently than in vitro. Thus, we found that structural plasticity can occur similarly in the adult neocortex in vivo as in the hippocampus in vitro, although it happens stringently in a smaller subset of spines.

Chronic epileptic foci in vitro in hippocampal slices from rats with the tetanus toxin epileptic syndrome

1989 ◽  
Vol 62 (2) ◽  
pp. 458-468 ◽  
Author(s):  
J. G. Jefferys
Keyword(s):  
Tetanus Toxin ◽  
Hippocampal Slices ◽  
Pyramidal Cells ◽  
Epileptic Activity ◽  
Field Potentials ◽  
Survival Times ◽  
Epileptic Syndrome ◽  
Epileptic Discharges

1. Minute doses of tetanus toxin were injected into the hippocampi of rats, under pentobarbitone anesthesia, to induce a chronic experimental epilepsy. The effects of this treatment were studied in vitro in hippocampal slices prepared 1-60 days after injection. 2. Epileptic activity was preserved in these slices in vitro, closely resembling that seen in vivo. Epileptiform afterdischarges were evoked by stimulation after survival times of greater than or equal to 3 days from injection. Spontaneous synchronous epileptic discharges were recorded from 7 days after injection. Both kinds of epileptiform activity were found with survival times up to 36 days but not beyond 44 days. This time course resembles the waxing and waning of the epileptic syndrome in vivo. 3. Two distinct types of spontaneous burst were seen. The first was a simple burst lasting 100-300 ms, reminiscent of the "interictal spike" of the clinical electroencephalogram. The second was much more prolonged, lasting several seconds. It consisted of a simple burst followed by a series of discrete afterbursts at 3-6/s and resembled the early stages of an epileptic seizure. Both types of burst were associated with slow field potentials that were positive at the cell-body layer. 4. Both the interictal and the seizure-like spontaneous epileptic discharges originated in the CA3b/c pyramidal cell region and propagated at 0.1-0.25 m/s along the cell layer toward the CA1 region. They occurred at very variable intervals, ranging from 20 s to 30 min. 5. Spontaneous epileptic bursts occurred in media containing 3 mM [K+]o to 5 mM in one-third of experiments during the period 1-4 wk after injection. Spontaneous bursts could be induced by increasing [K+]o to 5 mM in two-thirds of the remaining slices, which initially had produced evoked afterdischarges. 6. Intracellular recordings revealed that spontaneous field bursts were invariably associated with paroxysmal depolarization shifts (PDSs) and bursts of action potentials, suggesting that almost all the pyramidal cells in the region were recruited into the epileptic discharges. In some cells, smaller abnormal depolarizations were also seen; they were clearly larger than the spontaneous synaptic potentials but were not associated with field potentials. They may have been due to a more limited recruitment of pyramidal cells into partially synchronous bursts. 7. The tetanus toxin experimental epileptic syndrome differs from chronic models described previously in retaining in the hippocampal slice in vitro much of the spontaneous epileptic activity seen in vivo in the freely moving chronically epileptic rat.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Immunological unresponsiveness to native dextran B512 in young animals of dextran high responder strains is due to lack of Ig receptors expression. Evidence for a nonrandom expression of V-genes.

1978 ◽  
Vol 147 (3) ◽  
pp. 645-655 ◽  
Author(s):  
C Fernandez ◽  
G Möller
Keyword(s):  
High Responder ◽  
Age Difference ◽  
Protein Conjugates ◽  
Gene Translocation ◽  
Adult Mice ◽  
Gene Coding ◽  
V Genes ◽  
Expression Evidence ◽  
Young Mice

Young mice of dextran high responder strains were found to be complete nonresponders to the alpha-1-6 epitope of dextran during 30-40 days after birth. They also failed to respond to thymus-dependent dextran-protein conjugates. Cells from young and adult mice were activated equally well to polyclonal antibody synthesis by the polyclonal B-cell-activating property of dextran. There was no age difference in the immune response to haptens conjugated to dextran, indicating that dextran can function as an efficient carrier also in young mice. Unresponsiveness could not be attributed to suppressor T cells or to a suppressive environment in young animals, as shown by transfer experiments, in which living or irradiated cells from young and adult mice were admixed in various ways and transferred to irradiated recipients of different ages. Cells from young mice did not affect response of adult cells (and the reverse), nor did the age of the irradiated recipient influence the response. When lymphocytes from young and adult mice were polyclonally activated in vitro by lipopolysaccharide, only cells from young mice failed to synthesize antibodies against the alpha-1-6 epitope of dextran, although they produced antibodies of all other specificities tested for. It was concluded that young animals fail to express immunoglobulins directed against the alpha-1-6 epitope during the first 30-40 days after birth. Since the mice possess the VH gene coding for antibodies against this particular epitope, it was concluded that the timing of V gene expression is regulated during development, possibly at the V-C gene translocation level.

scholarly journals Epileptiform activity in mouse hippocampal slices induced by moderate changes in extracellular Mg2+, Ca2+, and K+

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Haiyu Liu ◽  
Sai Zhang ◽  
Liang Zhang
Keyword(s):  
Prolonged Exposure ◽  
Epileptiform Activity ◽  
Hippocampal Slices ◽  
Brain Slices ◽  
Experimental Investigations ◽  
High Frequency Stimulation ◽  
Field Potentials ◽  
Epileptiform Discharges ◽  
Frequency Stimulation

Abstract Background Rodent brain slices—particularly hippocampal slices—are widely used in experimental investigations of epileptiform activity. Oxygenated artificial cerebrospinal fluid (ACSF) is used to maintain slices in vitro. Physiological or standard ACSF containing 3–3.5 mM K+, 1–2 mM Mg2+, and 1–3 mM Ca2+ generally does not induce population epileptiform activity, which can be induced by ACSF with high K+ (8–10 mM), low Mg2+, or low Ca2+ alone or in combination. While low-Mg2+ ACSF without intentionally added Mg salt but with contaminating Mg2+ (≤ 50–80 µM) from other salts can induce robust epileptiform activity in slices, it is unclear whether such epileptiform activity can be achieved using ACSF with moderately decreased Mg2+. To explore this issue, we examined the effects of moderately modified (m)ACSF with 0.8 mM Mg2+, 1.3 mM Ca2+, and 5.7 mM K+ on induction of epileptiform discharges in mouse hippocampal slices. Results Hippocampal slices were prepared from young (21–28 days old), middle-aged (13–14 months old), and aged (24–26 months old) C57/BL6 mice. Conventional thin (0.4 mm) and thick (0.6 mm) slices were obtained using a vibratome and pretreated with mACSF at 35–36 °C for 1 h prior to recordings. During perfusion with mACSF at 35–36 °C, spontaneous or self-sustained epileptiform field potentials following high-frequency stimulation were frequently recorded in slices pretreated with mACSF but not in those without the pretreatment. Seizure-like ictal discharges were more common in thick slices than in thin slices. Conclusions Prolonged exposure to mACSF by pretreatment and subsequent perfusion can induce epileptiform field potentials in mouse hippocampal slices.

In Situ Localization of PPAR Gamma and Uncoupling Protein in Mouse Embryo Sections Using Digoxigenin-Labeled Riboprobes

1996 ◽  
Vol 54 ◽  
pp. 32-33
Author(s):  
C. Jennermann ◽  
S. A. Kliewer ◽  
D. C. Morris
Keyword(s):  
Alkaline Phosphatase ◽  
Room Temperature ◽  
Uncoupling Protein ◽  
Ppar Gamma ◽  
Nuclear Hormone Receptor ◽  
Full Length ◽  
Northern Analysis ◽  
Peroxisome Proliferator

Peroxisome proliferator-activated receptor gamma (PPARg) is a member of the nuclear hormone receptor superfamily and has been shown in vitro to regulate genes involved in lipid metabolism and adipocyte differentiation. By Northern analysis, we and other researchers have shown that expression of this receptor predominates in adipose tissue in adult mice, and appears first in whole-embryo mRNA at 13.5 days postconception. In situ hybridization was used to find out in which developing tissues PPARg is specifically expressed.Digoxigenin-labeled riboprobes were generated using the Genius™ 4 RNA Labeling Kit from Boehringer Mannheim. Full length PPAR gamma, obtained by PCR from mouse liver cDNA, was inserted into pBluescript SK and used as template for the transcription reaction. Probes of average size 200 base pairs were made by partial alkaline hydrolysis of the full length transcripts. The in situ hybridization assays were performed as described previously with some modifications. Frozen sections (10 μm thick) of day 18 mouse embryos were cut, fixed with 4% paraformaldehyde and acetylated with 0.25% acetic anhydride in 1.0M triethanolamine buffer. The sections were incubated for 2 hours at room temperature in pre-hybridization buffer, and were then hybridized with a probe concentration of 200μg per ml at 70° C, overnight in a humidified chamber. Following stringent washes in SSC buffers, the immunological detection steps were performed at room temperature. The alkaline phosphatase labeled, anti-digoxigenin antibody and detection buffers were purchased from Boehringer Mannheim. The sections were treated with a blocking buffer for one hour and incubated with antibody solution at a 1:5000 dilution for 2 hours, both at room temperature. Colored precipitate was formed by exposure to the alkaline phosphatase substrate nitrobluetetrazoliumchloride/ bromo-chloroindlylphosphate.

Evaluation of 99mTc-Label led Vitamin K4 as an Agent for Testicular Imaging

1991 ◽  
Vol 30 (01) ◽  
pp. 35-39 ◽  
Author(s):  
H. S. Durak ◽  
M. Kitapgi ◽  
B. E. Caner ◽  
R. Senekowitsch ◽  
M. T. Ercan
Keyword(s):  
Soft Tissue ◽  
Room Temperature ◽  
Normal Subjects ◽  
Left Testis ◽  
Wide Range ◽  
Activity Ratios ◽  
The Right ◽  
Radioactivity Levels

Vitamin K4 was labelled with 99mTc with an efficiency higher than 97%. The compound was stable up to 24 h at room temperature, and its biodistribution in NMRI mice indicated its in vivo stability. Blood radioactivity levels were high over a wide range. 10% of the injected activity remained in blood after 24 h. Excretion was mostly via kidneys. Only the liver and kidneys concentrated appreciable amounts of radioactivity. Testis/soft tissue ratios were 1.4 and 1.57 at 6 and 24 h, respectively. Testis/blood ratios were lower than 1. In vitro studies with mouse blood indicated that 33.9 ±9.6% of the radioactivity was associated with RBCs; it was washed out almost completely with saline. Protein binding was 28.7 ±6.3% as determined by TCA precipitation. Blood clearance of 99mTc-l<4 in normal subjects showed a slow decrease of radioactivity, reaching a plateau after 16 h at 20% of the injected activity. In scintigraphic images in men the testes could be well visualized. The right/left testis ratio was 1.08 ±0.13. Testis/soft tissue and testis/blood activity ratios were highest at 3 h. These ratios were higher than those obtained with pertechnetate at 20 min post injection.99mTc-l<4 appears to be a promising radiopharmaceutical for the scintigraphic visualization of testes.

Effect of Temperature on ADP-Induced Platelet Aggregation

1973 ◽  
Vol 29 (01) ◽  
pp. 183-189
Author(s):  
C. A Praga ◽  
E. M Pogliani
Keyword(s):  
Platelet Aggregation ◽  
Incubation Period ◽  
Room Temperature ◽  
Important Variable ◽  
Practical Significance ◽  
Effect Of Temperature ◽  
Induce Platelet Aggregation ◽  
Cuvette Holder ◽  
Exact Temperature

SummaryTemperature represents a very important variable in ADP-induced platelet aggregation.When low doses of ADP ( < 1 (μM) are used to induce platelet aggregation, the length of the incubation period of PRP in the cuvette holder of the aggregometer, thermostatted at 37° C, is very critical. Samples of the same PRP previously kept at room temperature, were incubated for increasing periods of time in the cuvette of the aggregometer before adding ADP, and a significant decrease of aggregation, proportional to the length of incubation, was observed. Stirring of the PRP during the incubation period made these changes more evident.To measure the exact temperature of the PRP during incubation in the aggre- gometer, a thermocouple device was used. While the temperature of the cuvette holder was stable at 37° C, the PRP temperature itself increased exponentially, taking about ten minutes from the beginning of the incubation to reach the value of 37° C. The above results have a practical significance in the reproducibility of the platelet aggregation test in vitro and acquire particular value when the effect of inhibitors of ADP induced platelet aggregation is studied.Experiments carried out with three anti-aggregating agents (acetyl salicyclic acid, dipyridamole and metergoline) have shown that the incubation conditions which influence both the effect of the drugs on platelets and the ADP breakdown in plasma must be strictly controlled.

Sign in / Sign up

Export Citation Format

Share Document