Is electrical coupling involved in the generation of posterior hypothalamic theta rhythm?

Data obtained in in vitro experiments and urethane anaesthetized animals have revealed that the mechanisms responsible for the generation of hippocampal cholinergic theta rhythm are specifically affected by the administration of broad spectrum gap junctions (GJs) blocker – carbenoxolone (CBX). The aim of this study was to examine the effect of GJs modulation on the production of posterior hypothalamic theta. Specifically, we were interested in evaluating whether CBX could attenuate the theta rhythm recorded from the supramammillary nucleus and posterior hypothalamic nuclei, in both in vitro and in vivo preparations. The data we obtained from in vitro and in vivo preparations demonstrated that the administration of CBX did not suppress cholinergically induced theta in posterior hypothalamic area (PHa) slices nor the theta rhythm observed in the PHa of urethane anaesthetized rats. Moreover, the application of trimethylamine, while very effective in the enhancement of hippocampal theta rhythm, did not produce any changes in theta oscillations observed in either in vitro or in vivo posterior hypothalamic area preparations. These data show that electrical coupling via GJs is not involved in theta rhythm generation in the PHa. Surprisingly, we observed a significant enhancement of theta activity in response to the carbenoxolone administration in both in vitro and in vivo PHa preparations. The theta rhythm enhancement detected in those experiments was attenuated by the application of spironolactone (mineralocorticoid receptors antagonist). We suggest that the observed excitatory effects of CBX on posterior hypothalamic oscillatory activity in the theta band could be mediated by mineralocorticoid receptors.     

Is the dentate gyrus an independent generator of in vitro recorded theta rhythm?

In this study we investigated the ability of the dentate gyrus to independently generate cholinergically induced theta rhythm in vitro. Two different experiments were performed. In Experiment I, new laminar profiles of theta phase, amplitude, and current sources and sinks were constructed. In this experiment a gradual phase shift of theta waves in the CA1 stratum radiatum was observed. Simultaneously, two amplitude maxima were detected: the first in the CA1 stratum oriens, and the second in the CA1 stratum lacunosum-moleculare. Moreover, during the positive peak of theta in the CA1 stratum oriens, two large sinks were observed: the first localized in the stratum oriens and the second in the stratum lacunosum-moleculare. In Experiment II the EEG activity of three different transected hippocampal slices (CA1 transected slices, CA3c transected slices, and DG transected slices) was recorded. It was demonstrated that the dentate gyrus granular cell body layer was not able to independently produce in vitro theta rhythm. Data obtained in both experiments provide strong evidence that in cholinegically treated hippocampal formation maintained in vitro there is no independent generator of theta rhythm in the region of the dentate gyrus granular cell layer.     

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.     

Firing cell repertoire during carbachol-induced theta rhythm in rat hippocampal formation slices

One hundred and seven cells were recorded extracellularly in hippocampal formation (HPC) slices during carbachol-induced theta. The data obtained provided evidence of a population of HPC neurons which, when activated cholinergically, participate in the generation of in vitro theta. The activity patterns of in vitro recorded theta-related cells were shown to be similar to those of theta-related cells recorded in vivo and cells recorded in vitro during cholinergically induced theta, and non-theta intervals were successfully classified according to previously developed criteria for in vivo recorded theta-related cells. The current in vitro experiments showed that, in addition to theta-on and theta-off cells, the HPC contained cells that were probably involved in programming the appearance and duration of theta epochs and the intervals between theta epochs. These novel types of cells were termed 'gating cells'.     

Intrinsic oscillations in CA3 hippocampal pyramids: Physiological relevance to theta rhythm generation

The theta rhythm is the most remarkable hippocampal activity correlated with various physiological and behavioral phenomena. Although analyzed by numerous investigators, during the last five decades, the mechanisms that lead to its generation still remain reason of debate. In the present report it is shown that hippocampal neurons, recorded from juvenile and adult slices, are endowed with intrinsic properties that allow the generation of a steady oscillatory activity. The frequency of this rhythmic ongoing process is highly sensitive to the level of the membrane potential, reaching values in the theta range, up to 6 Hz, for more depolarized values of membrane potential. Membrane potential oscillations are unmasked by loading pyramidal neurons with intracellular cesium, are sodium-independent, and are generated by the sequential activation of calcium and potassium conductances. Finally, like theta rhythm, regularly occurring membrane potential oscillations can be detected since postnatal day 10 and their frequency increases with age, i.e., during the following 2 weeks. Hippocampus 8:666–679, 1998. © 1998 Wiley-Liss, Inc.     

The effect of carbenoxolone on hippocampal formation theta rhythm in rats: In vitro and in vivo approaches

The role of gap junction (GJ) coupling in the generation of hippocampal formation theta rhythm was investigated in vitro, with use of brain slices, and in vivo, with use of urethane anesthetized rats. Carbenoxolone, the succinyl ester of glycyrrhetinic acid, and GJ blocker reversibly abolished hippocampal formation theta rhythm recorded in slice preparations and urethane anesthetized rats. The present study yielded novel data which demonstrated that the pattern of delay in blockage of theta rhythm after carbenoxolone treatment, and the pattern of theta recovery after administration of this agent, require a specific time period (2–3 h for delay and 8–12 h for recovery), one that can be demonstrated using different experimental protocols.     

The effect of atropine administered in the medial septum or hippocampus on high- and low-frequency theta rhythms in the hippocampus of urethane anesthetized rats

Cholinergic mechanisms are critical for the generation of hippocampal theta rhythm. Cholinergic innervation of the hippocampus originates from the medial septum (MS) and cholinergic receptors are expressed in both the MS and hippocampus. In this study, we compared the effects of the muscarinic receptor antagonist atropine in the MS and the hippocampus on theta generation. Hippocampal theta rhythm was elicited by electrical stimulation of the pontine reticular formation using series of stimuli with varying intensities. Atropine was administered either systemically (50 mg/kg i.p.) or locally in the MS (microdialysis; 25 and 75 mM for 30 or 90 min) or in the hippocampus on one side (microinjection; 20 or 40 ug). The relative power at the peak theta frequency was calculated and averaged over episodes of low-intensity and high-intensity stimulations. We found that atropine drastically reduced theta rhythmic synchronization when injected in either location. After MS administration of atropine, however, high-frequency theta elicited by high-intensity stimuli was more resistant (58% and 67% decrease after 25 mM and 75 mM atropine, respectively) than slow theta elicited by low-intensity stimuli (86% and 91% decrease). There was no significant difference between the powers of the two oscillations after hippocampal injections (70-75% decrease). We conclude that the theta suppressing effect of atropine involves both hippocampal and septal mechanisms and that low-frequency theta as compared with fast theta rhythm is more sensitive to muscarinic acetylcholine receptor antagonism in the MS but not in the hippocampus.     

Development of NMDA-induced theta rhythm in hippocampal formation slices

During the past 20 years experimental evidence has accumulated demonstrating that the appearance of theta rhythm requires a certain level of excitation of local neuronal networks. In this study we extended our earlier in vitro observations concerning the involvement of cholinergic and GABAergic neurotransmission in hippocampal theta production. Specifically, we investigated whether the hippocampal neuronal network is capable of generating theta oscillations in the presence of N-methyl-d-aspartic acid (NMDA) in a brain slice preparation. To answer this question, the effect of different concentrations of NMDA (Experiment I) and the effect of interaction between NMDA and GABA_A/_B agonists and antagonists on field potentials recorded in the CA3c region of hippocampal formation (HPC) slice preparations (Experiment II) was examined. We demonstrated for the first time that apart from the epileptiform activity recorded in almost all series of Experiments I and II, only the perfusion of HPC slices with NMDA in doses of 30 and 50 μM, as well as the perfusion of HPC slices with NMDA and GABA_B agonist baclofen (50 μM NMDA + 50 μM BACL), resulted in the appearance of individual theta epochs. The best synchronized theta oscillations obtained after administration of 50 μM NMDA + 50 μM BACL resembled theta activity induced by a bath perfusion of 50 μM carbachol. In light of the obtained results we conclude that besides the cholinergic and GABAergic input, NMDA glutamatergic drive is also important for the appearance of theta oscillations in HPC in vitro.     

Septohippocampal GABAergic neurons mediate the altered behaviors induced by n‐methyl‐D‐aspartate receptor antagonists

We hypothesize that selective lesion of the septohippocampal GABAergic neurons suppresses the altered behaviors induced by an N‐methyl‐D ‐aspartate (NMDA) receptor antagonist, ketamine or MK‐801. In addition, we hypothesize that septohippocampal GABAergic neurons generate an atropine‐resistant theta rhythm that coexists with an atropine‐sensitive theta rhythm in the hippocampus. Infusion of orexin‐saporin (ore‐SAP) into the medial septal area decreased parvalbumin‐immunoreactive (GABAergic) neurons by ～80%, without significantly affecting choline‐acetyltransferase‐immunoreactive (cholinergic) neurons. The theta rhythm during walking, or the immobility‐associated theta induced by pilocarpine, was not different between ore‐SAP and sham‐lesion rats. Walking theta was, however, more disrupted by atropine sulfate in ore‐SAP than in sham‐lesion rats. MK‐801 (0.5 mg/kg i.p.) induced hyperlocomotion associated with an increase in frequency, but not power, of the hippocampal theta in both ore‐SAP and sham‐lesion rats. However, MK‐801 induced an increase in 71–100 Hz gamma waves in sham‐lesion but not ore‐SAP lesion rats. In sham‐lesion rats, MK‐801 induced an increase in locomotion and an impairment of prepulse inhibition (PPI), and ketamine (3 mg/kg s.c.) induced a loss of gating of hippocampal auditory evoked potentials. MK‐801‐induced behavioral hyperlocomotion and PPI impairment, and ketamine‐induced auditory gating deficit were reduced in ore‐SAP rats as compared to sham‐lesion rats. During baseline without drugs, locomotion and auditory gating were not different between ore‐SAP and sham‐lesion rats, and PPI was slightly but significantly increased in ore‐SAP as compared with sham lesion rats. It is concluded that septohippocampal GABAergic neurons are important for the expression of hyperactive and psychotic symptoms an enhanced hippocampal gamma activity induced by ketamine and MK‐801, and for generating an atropine‐resistant theta. Selective suppression of septohippocampal GABAergic activity is suggested to be an effective treatment of some symptoms of schizophrenia. © 2012 Wiley Periodicals, Inc.     

Orexinergic theta rhythm in the rat hippocampal formation: In vitro and in vivo findings

Previous in vivo data suggested that orexin neuropeptides (ORX_A and ORX_B) synthetized in hypothalamic neurons were involved in the mechanism of generation of the hippocampal formation theta rhythm. Surprisingly, this suggestion has never been directly proved by experiments using intraseptal or intrahippocampal administration of orexins. In this study, involving the use of in vitro hippocampal formation slices and in vivo model of anesthetized rat, we provide the first convergent electropharmacological evidence that in the presence of both ORX_A and ORX_B the hippocampal formation neuronal network is capable of producing oscillations in the theta band. This effect of orexin peptides was antagonized by selective blockers of orexin receptors (OX₁R and OX₂R), SB 334867 and TCS OX2 29, respectively. These results provide evidence for a novel, orexinergic mechanism responsible for the production of theta rhythm in the hippocampal formation neuronal network. © 2015 Wiley Periodicals, Inc.     

The effect of posterior hypothalamic injection of cholinergic agents on hippocampal formation theta in freely moving cat

The effect of intra-hypothalamic micro-injection of muscarinic (atropine sulphate, pirenzepine and gallamine) and nicotinic (hexamethonium) antagonists on spontaneous, sensory and electrically-induced hippocampal formation (HPC) theta EEG activity was investigated in the freely behaving cat. Administration of hexamethonium and gallamine failed to elicit a detectable effect on HPC theta activity. However, the injection of atropine sulphate and pirenzepine abolished the theta rhythm recorded from HPC. This effect was reversible. A substantial difference in the recovery time course between frequency versus amplitude (microV) and power (microV2) of hippocampal theta was observed. While theta frequency exhibited a rapid reappearance with a shallow slope, the power and amplitude showed a gradual recovery with a steeper slope. The injection of carbachol into posterior hypothalamus (PH) produced almost a continuous HPC theta with increased power. These results demonstrate that cholinergic (M1) receptors localised in the posterior area of the hypothalamus are engaged in mechanisms responsible for generating hippocampal theta oscillations in the freely behaving cat. The contribution of posterior hypothalamic region to HPC theta frequency and amplitude is discussed.     

Obituary: Cornelius H. Vanderwolf

C.H. Vanderwolf described motor correlates of hippocampal theta oscillations and uncovered two broad classes: atropine‐sensitive and atropine‐resistant rhythm with likely different behavioral and cognitive significance. © 2015 Wiley Periodicals, Inc.     

Human cerebrospinal fluid promotes spontaneous gamma oscillations in the hippocampus in vitro

Gamma oscillations (30–80 Hz) are fast network activity patterns frequently linked to cognition. They are commonly studied in hippocampal brain slices in vitro, where they can be evoked via pharmacological activation of various receptor families. One limitation of this approach is that neuronal activity is studied in a highly artificial extracellular fluid environment, as provided by artificial cerebrospinal fluid (aCSF). Here, we examine the influence of human cerebrospinal fluid (hCSF) on kainate‐evoked and spontaneous gamma oscillations in mouse hippocampus. We show that hCSF, as compared to aCSF of matched electrolyte and glucose composition, increases the power of kainate‐evoked gamma oscillations and induces spontaneous gamma activity in areas CA3 and CA1 that is reversed by washout. Bath application of atropine entirely abolished hCSF‐induced gamma oscillations, indicating critical contribution from muscarinic acetylcholine receptor‐mediated signaling. In separate whole‐cell patch clamp recordings from rat hippocampus, hCSF increased theta resonance frequency and strength in pyramidal cells along with enhancement of h‐current (I_h) amplitude. We found no evidence of intrinsic gamma frequency resonance at baseline (aCSF) among fast‐spiking interneurons, and this was not altered by hCSF. However, hCSF increased the excitability of fast‐spiking interneurons, which likely contributed to gamma rhythmogenesis. Our findings show that hCSF promotes network gamma oscillations in the hippocampus in vitro and suggest that neuromodulators distributed in CSF could have significant influence on neuronal network activity in vivo.     

In vitro activation of the medial septum-diagonal band complex generates atropine-sensitive and atropine-resistant hippocampal theta rhythm: an investigation using a complete septohippocampal preparation

The medial septum and diagonal band complex (MS-DB) is believed to play a key role in generating theta oscillations in the hippocampus, a phenomenon critical for learning and memory. Although the importance of the MS-DB in hippocampal theta rhythm generation is generally accepted, it remains to be determined whether the MS-DB alone can generate hippocampal oscillations or is only a transducer of rhythmic activity from other brain areas. Secondly, it is known that hippocampal theta rhythm can be separated into an atropine-sensitive and insensitive component. However, it remains to be established if the MS-DB can generate both types of rhythm. To answer these questions, we used a new in vitro rat septohippocampal preparation placed in a hermetically separated two side recording chamber. We showed that carbachol activation of the MS-DB generated large theta oscillations in the CA1 and CA3 regions of the hippocampus. These oscillations were blocked by applying either the GABA(A) receptor antagonist bicuculline or the AMPA/kainate antagonist DNQX to the hippocampus. Interestingly, the application of the muscarinic receptor antagonist atropine produced only a partial decrease in the amplitude, without modification of the frequency, of theta. These results show for the first time, that upon optimal excitation, the MS-DB alone is able to generate hippocampal oscillations in the theta frequency band. Moreover, these MS-DB generated theta oscillations are mediated by muscarinic and nonmuscarinic receptors and have a pharmacological profile similar to theta rhythm observed in awake animals.     

In vivo intrahippocampal microinfusion of carbachol and bicuculline induces theta-like oscillations in the septally deafferented hippocampus.

In their laboratory the authors have previously demonstrated that hippocampal slices could be induced to generate trains of "theta-like" oscillations by whole-bath perfusions of carbachol. Until recently, it has not been possible to generate similar activity in the septally deafferented hippocampus of an otherwise intact brain by microinfusions of carbachol. This study presents a full report of the first demonstration of a theta-like oscillation in the in vivo, septally deafferented hippocampal formation. Rats were anesthetized with urethane and implanted with microinfusion cannulae in the region of the medial septum/vertical limb of the diagonal band of Broca (MS/vDBB) and at single or multiple sites in the stratum moleculare of the fascia dentata. The MS/vDBB was microinfused with procaine hydrochloride to produce a reversible suppression lasting for approximately 20 minutes. Intrahippocampal microinfusions of carbachol or bicuculline alone (in the postprocaine condition of the MS/vDBB) failed to produce any theta-like oscillations. The combination of carbachol and bicuculline produced trains of theta-like oscillations during suppression of the MS/vDBB very similar to those seen in the slice preparations. The oscillations were blocked by intravenous administration of atropine sulfate, and they had the same depth profile as that of theta. Theta-on cells were shown to discharge in rhythmic bursts in synchrony with the oscillations. Thus, it would appear that the essential nature of the medial septal input to the hippocampal formation, for the generation of theta field activity in the intact brain, consists of a critical balance between cholinergic and GABAergic circuitry.     

Spatio‐temporal dynamics of theta oscillations in hippocampal–entorhinal slices

Theta oscillations (4–12 Hz) are associated with learning and memory and are found in the hippocampus and the entorhinal cortex (EC). The spatio‐temporal organization of rhythmic activity in the hippocampal–EC complex was investigated in vitro. The voltage sensitive absorption dye NK3630 was used to record the changes in aggregated membrane voltage simultaneously from the neuronal networks involved. Oscillatory activity at 7.0 Hz (range, 5.8–8.2) was induced in the slice with the muscarinic agonist carbachol (75–100 μM) in the presence of bicuculline (5 μM). Time relations between all recording sites were analyzed using cross‐correlation functions which revealed systematic phase shifts in the theta oscillation recorded from the different entorhinal and hippocampal subregions. These phase shifts could be interpreted as propagation delays. The oscillation propagates over the slice in a characteristic spatio‐temporal sequence, where the entorhinal cortex leads, followed by the subiculum and then the dentate gyrus (DG), to finally reach the CA3 and the CA1 area. The delay from dentate gyrus to the CA3 area was 12.4 ± 1.1 ms (mean ± s.e.m.) and from the CA3 to the CA1 region it was 10.9 ± 1.9 ms. The propagation delays between the hippocampal subregions resemble the latencies of electrically evoked responses in the same subregions. Removing the entorhinal cortex from the slice changed the spatiotemporal pattern into a more clustered pattern with higher local synchrony. We conclude that in the slice, carbachol‐induced theta oscillations are initiated in the entorhinal cortex. The EC could serve to control the information flow through the neuronal network in the subregions of the hippocampus by synchronizing and/or entraining their responses to external inputs. © 2009 Wiley‐Liss, Inc.     

Passive rotation‐induced theta rhythm and orientation homeostasis response

Rhythmic oscillatory activities at the theta frequency (3–12 Hz) have long attracted attention, as they have been implicated in diverse brain functions. There are two kinds of hippocampal theta rhythms: Type 1 is an atropine‐resistant noncholinergic theta rhythm, and Type 2 is an atropine‐sensitive cholinergic theta rhythm. However, it has not yet been determined whether the theta rhythm generated during passive whole‐body rotation is of Type 1 or 2. To clarify this issue, we investigated passive whole‐body rotation‐induced theta rhythm using C57BL/6J normal and atropine‐treated mice. The results demonstrated that atropine [50 mg/kg, intraperitoneally (i.p.)], a cholinergic antagonist, abolished the theta rhythm generated during passive whole‐body rotation. Therefore, the passive whole‐body rotation‐induced theta rhythm is an atropine‐sensitive Type 2 theta rhythm. In addition, we found that blocking cholinergic receptors using atropine resulted in the loss of the orientation homeostasis response, which is a circling behavior in the direction opposite to that of the rotating circular treadmill that is generated to maintain a constant orientation. These results suggest that atropine‐sensitive Type 2 theta rhythm can be generated by a passive rotation‐induced vestibular sensory signal and may be necessary for spatial orientation homeostasis response behavior. Synapse 64:409–415, 2010. © 2009 Wiley‐Liss, Inc.     

Regular theta‐firing neurons in the nucleus incertus during sustained hippocampal activation

This paper describes the existence of theta‐coupled neuronal activity in the nucleus incertus (NI). Theta rhythm is relevant for cognitive processes such as spatial navigation and memory processing, and can be recorded in a number of structures related to the hippocampal activation including the NI. Strong evidence supports the role of this tegmental nucleus in neural circuits integrating behavioural activation with the hippocampal theta rhythm. Theta oscillations have been recorded in the local field potential of the NI, highly coupled to the hippocampal waves, although no rhythmical activity has been reported in neurons of this nucleus. The present work analyses the neuronal activity in the NI in conditions leading to sustained hippocampal theta in the urethane‐anaesthetised rat, in order to test whether such activation elicits a differential firing pattern. Wavelet analysis has been used to better define the neuronal activity already described in the nucleus, i.e., non‐rhythmical neurons firing at theta frequency (type I neurons) and fast‐firing rhythmical neurons (type II). However, the most remarkable finding was that sustained stimulation activated regular‐theta neurons (type III), which were almost silent in baseline conditions and have not previously been reported. Thus, we describe the electrophysiological properties of type III neurons, focusing on their coupling to the hippocampal theta. Their spike rate, regularity and phase locking to the oscillations increased at the beginning of the stimulation, suggesting a role in the activation or reset of the oscillation. Further research is needed to address the specific contribution of these neurons to the entire circuit.     

Posterior hypothalamic GABAergic mediation of hippocampal theta in the cat

The effect of posterior hypothalamic (PH) microinjections of GABA_A and GABA_B agonists (muscimol and baclophen, respectively) and antagonists (bicuculline and 2-OH saclophen) on spontaneous, sensory and electrically induced hippocampal formation (HPC) theta EEG activity was investigated in freely behaving cats. Administration of GABAergic agonists abolished the theta rhythm recorded from HPC. This effect was reversible. A substantial difference in the recovery time course between frequency versus amplitude and power of hippocampal theta was observed. While theta frequency exhibited a rapid reappearance with a shallow slope, the power and amplitude showed a gradual recovery with a steeper slope. The PH injection of GABAergic antagonists produced HPC theta with increased power. These results demonstrate that both types of GABAergic receptors localized in PH are engaged in mechanisms responsible for generating hippocampal theta oscillations in freely behaving cats. The present study provides additional support for the essential difference between rats and cats in the programming of HPC theta amplitude and frequency. While the PH in rats is involved in programming the frequency of theta rhythm, the same region in cats mainly determines theta amplitude.     

Recognition memory and theta–gamma interactions in the hippocampus

Neuronal oscillations and cross‐frequency interactions in the rat hippocampus relate in important ways to memory processes and serve as a model for studying oscillatory activity in cognition more broadly. We report here that hippocampal synchrony (CA3–CA1 coherence) increased markedly in the low gamma range as rats were exploring novel objects, particularly those for which the rat subsequently showed good memory. The gamma synchrony varied across phases of the theta rhythm such that coherence was highest at the falling slope and trough of the theta wave. Further, the shape of the theta wave was more asymmetric and elongated at the falling slope during exploration of objects for which the rat subsequently showed good memory as compared with objects for which the rat subsequently showed poor memory. The results showed a strong association between event‐related gamma synchrony in rat hippocampus and memory encoding for novel objects. In addition, a novel potential mechanism of cross‐frequency interactions was observed whereby dynamic alterations in the shape of theta wave related to memory in correspondence with the strength of gamma synchrony. These findings add to our understanding of how theta and gamma oscillations interact in the hippocampus in the service of memory. © 2013 Wiley Periodicals, Inc.