Jan Konopacki

Carbachol-induced EEG ‘theta’ activity in hippocampal brain slices

Application of the cholinergic agonist carbachol (50 microM) produced theta-like rhythmical waveforms, recorded in the stratum moleculare of the dentate gyrus. Atropine sulfate (50 microM) antagonized the carbachol-induced theta-like activity, consistent with this action of atropine in vivo. These results provide the first direct evidence that hippocampal neurons are capable of producing synchronized slow-wave activity when isolated from pulsed rhythmic inputs of the medial septum and other brain regions.

Intracellular records of carbachol-induced theta rhythm in hippocampal slices ☆

Intracellular recordings were made in the CA1, CA3 and dentate cell layers prior to, during and after the bath perfusion of 50 microM carbachol on hippocampal slices. Fifty-six percent of the cells in this sample were termed theta (theta)-related, i.e., they exhibited membrane potential oscillations of 5-28 mV and rhythmic spike discharges related to the carbachol-induced extracellular theta-rhythm. The remaining 44% of the cells did not show the above relationships to the extracellular theta-rhythm. Carbachol produced an overall depolarization in all cells, in the range of 10-20 mV. These results demonstrated the cellular basis of carbachol-induced theta in hippocampal slices. This preparation will be a valuable model for studying cellular mechanisms and network properties underlying electroencephalographic activity.

In vivo intracellular correlates of hippocampal formation theta-on and theta-off cells

Using urethane-anesthetized rat, intracellular recordings were made in hippocampal formation cells classified according to previously established criteria as either theta-on or theta-off, in order to further define the electrophysiological characteristics of these cells. Four cells classified as phasic theta-off cells had short duration spikes (less than 1 ms), high input resistances (54-61 M omega) and large fast afterhyperpolarizations (6-10 mV), thus sharing some of the properties of identified hippocampal interneurons. Phasic theta-off cells also exhibited rhythmic membrane potential oscillations (MPOs) ranging from 4 to 10 mV in amplitude during the simultaneous occurrence of extracellular theta field activity, but not during the occurrence of large amplitude irregular field activity (LIA). The MPOs of phasic theta-off cells were the same frequency as and were highly coherent with the extracellular theta field activity. In all four phasic theta-off cells the positive peak of the MPO was in phase with the positive peak of the local theta field activity. At the onset of extracellular theta field activity above 4-5 Hz, the membrane potentials of phasic theta-off cells showed a 5-10-mV hyperpolarizing shift, accompanied by MPOs without spike discharges. As theta frequency slowed down there was a return to baseline membrane potential levels and spike discharges occurred near the positive peak of the MPOs. The seven cells classified as phasic theta-on had longer duration spikes (greater than 1 ms), lower input resistances (22-36 M omega) and small (approx. 1.0 mV) fast afterhyperpolarizations, thus sharing some of the properties of hippocampal projection cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbachol-induced EEG ‘theta’ in hippocampal formation slices: evidence for a third generator of theta in CA3c area

The topography of carbachol-induced EEG theta activity was studied using the hippocampal formation slice preparation. Systematic tracking with electrodes exhibited two amplitude maxima of cholinergic-induced theta, one located in the stratum oriens of the CA1 pyramidal cells and the other in a region of CA3c pyramidal neurons. In addition, mapping experiments demonstrated EEG theta in the CA3a and CA3b cell body layers, but not in the subicular and parasubicular regions, or the ventral blade of the dentate gyrus. Furthermore, transected slice (trans-slice) preparations used in the present study revealed that the CA3c region could generate carbachol-induced theta independently of CA1 and dentate gyrus generator zones and conversely, CA1 and dentate gyrus areas were capable of generating cholinergic-induced theta rhythm independently of the CA3c region. These results provide strong evidence for 3 independent, anatomically separated generators of theta: one located in the stratum oriens of CA1 neurons, a second in the stratum moleculare of the dentate gyrus and a third one in the region of Ca3c cells. In addition, the results support previous in vivo suggestions that theta rhythm can be either elicited or blocked by cholinergic agents acting on sites within the hippocampal formation.

Theta-like activity in hippocampal formation slices: cholinergic-GABAergic interaction

Brain slice preparations obtained from the rat were used to study cholinergic GABAergic interaction in mechanisms responsible for production of theta-like activity in hippocampal slices maintained in vitro. Bicuculline, a GABA-A antagonist, applied at 25 microM facilitated the effect of low concentration carbachol (25 microM) in inducing theta-like oscillations. At 100 microM, bicuculline increased the amplitude of carbachol-induced theta-like slow waves. This carbachol-bicuculline induced field potential was antagonized by a muscarinic blocker, atropine sulphate, and a GABA-A agonist, muscimol. These results provide in vitro evidence for cholinergic-GABAergic interaction in the production of hippocampal theta-like slow waves.

Evidence that activation of in vitro hippocampal θ rhythm only involves muscarinic receptors

Abstract The present study was conducted for two purposes: the first was to evaluate whether activation of nicotinic receptors in the hippocampal formation in vitro (slice) preparation was capable of producing type 2 (atropine-sensitive) θ rhythm. The cholinergic nature and involvement of muscarinic receptors in this type of θ has been previously well documented. The second purpose was to determine whether perfusion of a number of (other) putative neurotransmitters shown to be present in the hippocampal formation could elicit type 1 (atropine-resistant) θ in the slice preparation. Further experiments were conducted to determine if these agents interacted in any manner with cholinergically-induced type 2 θ. Electroencephalic (EEG) θ activity was not induced by nicotine, providing evidence for an exclusive muscarinic receptor involvement in this cholinergically-induced type 2 θ. In addition, θ activity was not elicited by the application of γ-aminobutyric acid (GABA), glutamate, norepinephrine, dopamine or serotonin. The application of any of these agents did not significantly alter the production of cholinergically-induced θ. These results suggest that type 1 θ originates in regions extrinsic to the hippocampus, or is the result of the interaction of several neurotransmitters on different receptors.

Theta-like Activity in the Limbic Cortex In Vitro

The generation of EEG theta rhythm in the mammalian limbic cortex is a prime example of rhythmic activity that involves central mechanisms of oscillations and synchrony. This EEG pattern has been extensively studied since 1938, when Jung and Kornmuller (28) (Eine methodik der ableitung lokalisierter potential schwankingen aus subcorticalen hirnyebieten, Arch. Psychiat. Neruenkr. 109 (1938) 1-30) demonstrated the first theta recordings in the hippocampal formation of rabbits. In 1986 we demonstrated for the first time that bath perfusion of hippocampal slices with the cholinergic agonist, carbachol, resulted in theta-like oscillations. Since this initial demonstration of in vitro theta-like activity, we have carried out a number of experiments in an attempt to answer the following question: what are the similarities between cholinergic-induced in vitro theta-like activity and theta rhythm which naturally occurs in the in vivo preparation. Thus far, our studies have provided strong evidence that theta-like activity recorded in vitro shares many of the physiological and pharmacological properties of theta rhythm observed in vivo.

Spatiotemporal Characterization of mTOR Kinase Activity Following Kainic Acid Induced Status Epilepticus and Analysis of Rat Brain Response to Chronic Rapamycin Treatment

Mammalian target of rapamycin (mTOR) is a protein kinase that senses nutrient availability, trophic factors support, cellular energy level, cellular stress, and neurotransmitters and adjusts cellular metabolism accordingly. Adequate mTOR activity is needed for development as well as proper physiology of mature neurons. Consequently, changes in mTOR activity are often observed in neuropathology. Recently, several groups reported that seizures increase mammalian target of rapamycin (mTOR) kinase activity, and such increased activity in genetic models can contribute to spontaneous seizures. However, the current knowledge about the spatiotemporal pattern of mTOR activation induced by proconvulsive agents is rather rudimentary. Also consequences of insufficient mTOR activity on a status epilepticus are poorly understood. Here, we systematically investigated these two issues. We showed that mTOR signaling was activated by kainic acid (KA)-induced status epilepticus through several brain areas, including the hippocampus and cortex as well as revealed two waves of mTOR activation: an early wave (2 h) that occurs in neurons and a late wave that predominantly occurs in astrocytes. Unexpectedly, we found that pretreatment with rapamycin, a potent mTOR inhibitor, gradually (i) sensitized animals to KA treatment and (ii) induced gross anatomical changes in the brain.

Theta-like activity in hippocampal formation slices : the effect of strong disinhibition of GABAA and GABAB receptors

The involvement of GABAA and GABAB receptors in neural mechanisms responsible for the production of theta rhythms in hippocampal formation (HPC) slices is addressed in the present study. In a number of papers published in the last decade, we have demonstrated that theta-like activity can be successfully recorded in the limbic cortex maintained in vitro when the cholinergic agonists, acetylcholine, carbachol or muscarine, were added to the bath. Recently, we have also shown a strong GABAA modulation of the cholinergic-induced in vitro theta-like activity. This study presents a report of the first demonstration of in vitro theta-like field responses induced a consequence of simultaneously inhibiting hippocampal GABAA and GABAB receptors. HPC slices (350 microns) were maintained in a gas-liquid interface chamber (35 degrees C). Theta-like activity was induced in the presence of bath perfusion of bicuculline (GABAA antagonist) and 2-hydroxysaclophen (GABAB antagonist). This in vitro induced field response was antagonized both by muscimol (GABAA agonist) and baclophen (GABAB agonist). In addition, the experiments presented here revealed that bicuculline/2-hydroxysaclophen-induced in vitro theta-like activity also had a strong cholinergic M1 involvement: it was abolished by hemicholinium-3 (choline transport blocker) and pirenzepine (specific antagonist of M1 receptor), but not by gallamine (specific antagonist of M2 receptor). The results of the present study provided further evidence for a strong GABAergic/cholinergic interaction in the neural mechanism responsible for production of theta-like activity in the hippocampal formation slices.

Electrical coupling underlies theta oscillations recorded in hippocampal formation slices

The role of gap junction coupling in generation of carbachol-induced theta-like activity (TLA) in hippocampal formation (HPC) slices was investigated in this study. Two gap junction (GJ) blockers, carbenoxolone (100 microM) and quinine (100 microM), were tested. Both GJ blockers abolished cholinergically induced theta-like activity and related cell discharges. Abolishing effects were observed after 40-45 min of drug perfusion. These effects were found to be slowly and partially reversible. Our results provide evidence for the contribution of gap junction communication in mechanisms of neural synchrony, underlying the production of theta oscillations in limbic cortex maintained in vitro.