NMDA-glutamatergic activation of the ventral tegmental area induces hippocampal theta rhythm in anesthetized rats.

Glutamate afferents reaching the ventral tegmental area (VTA) affect dopamine (DA) cells in this structure probably mainly via NMDA receptors. VTA appears to be one of the structures involved in regulation of hippocampal theta rhythm, and this work aimed at assessing the role of glutamatergic activation of the VTA in the theta regulation. Male Wistar rats (n=17) were divided into groups, each receiving intra-VTA microinjection (0.5 µl) of either solvent (water), glutamatergic NMDA agonist (0.2 µg) or antagonist (MK-801, 3.0 µg). Changes in local field potential were assessed on the basis of peak power (Pmax) and corresponding peak frequency (Fmax) for the delta (0.5-3 Hz) and theta (3-6 Hz) bands. NMDA microinjection evoked long-lasting hippocampal theta. The rhythm appeared with a latency of ca. 12 min post-injection and lasted for over 30 min; Pmax in this band was significantly increased for 50 min, while simultaneously Pmax in the delta band remained lower than in control conditions. Theta Fmax and delta Fmax were increased in almost entire post-injection period (by 0.3-0.5 Hz and 0.3-0.7 Hz, respectively). MK-801 depressed the sensory-evoked theta: tail pinch could not induce theta for 30 min after the injection; Pmax significantly decreased in the theta band and at the same time it increased in the delta band. Theta Fmax decreased 10 and 20 min post injection (by 0.4-0.5 Hz) and delta Fmax decreased in almost entire post injection period (by 0.3-0.7 Hz). NMDA injection generates theta rhythm probably through stimulation of dopaminergic activity within the VTA.

Locomotor response to novelty correlates with differences in number and morphology of hypothalamic tyrosine hydroxylase positive cells in rats.

Individual differences in the intensity of locomotor response to a new environment (exploratory reaction) are generally used as a model to study individual vulnerability to stress and drug addiction. In the present work we studied the number, distribution and morphology of the hypothalamic cells expressing tyrosine hydroxylase (TH+ cells) (immunohistochemical and immunofluorescent staining) in male Wistar rats divided based on high (HR), midline (MR) or low (LR) locomotor activity in response to novelty. Morphology and total number of TH+ cells were analyzed for A11-A15 dopaminergic groups. We found correlation between the total number of hypothalamic TH+ cells in the whole A11-A15 area and the locomotor activity. The differences were most pronounced in some of the hypothalamic nuclei, i.e. in the rostro-caudal extension of the A11, A12 and A14 structures, where the HR rats had a significantly higher number of TH+ cells in comparison to the MR and LR rats. Morphology analysis of TH+ cells showed HR/MR/LR differences in single cell area and perimeter and, to a lesser extent, in the other morphometric parameters such as length of the major and minor axes, or circularity factor. The results suggest that the behavioral traits which characterize the HR animals and are correlated with increased susceptibility to stress and propensity to develop drug addictions can be determined by the number, distribution, activity and perhaps the morphology of the cells in the dopaminergic systems.

Hippocampal theta rhythm induced by rostral pontine nucleus stimulation in the conditions of pedunculopontine tegmental nucleus inactivation.

Theta rhythm in rat hippocampus occurs during cortical activation in different forms of waking as well as during paradoxical phase of sleep. The multi-level regulatory system of theta, based mainly on cholinergic transmission, includes structures from the forebrain to the medulla. Among them the most important are two reticular nuclei: the pedunculopontine tegmental nucleus (PPN) and rostral pontine tegmental nucleus (RPO). Functional relations between these two nuclei are still unidentified. It is known that cholinergic stimulation of these nuclei with carbachol leads to induction of theta in the hippocampus. Electrical stimulation has the same effect but only when applied to the RPO. In our experiments, performed on urethanized rats, each of these two methods was applied to the RPO with the PPN being inactivated in the contralateral hemisphere. We found that inactivation of the PPN does not suppress theta induced with carbachol microinjection into the RPO, but completely blocks theta induction with electrical stimulation of the RPO. The results suggest the important role of the PPN in theta rhythm generation from brainstem level, depending on the method of theta rhythm induction, i.e. cholinergic or electric stimulation of the RPO.

Dopaminergic transmission in the midbrain ventral tegmental area in the induction of hippocampal theta rhythm.

Hippocampal rhythmic slow activity (RSA, theta) is regulated by many brainstem structures, including the midbrain ventral tegmental area (VTA). This work aimed at assessing the role of the dopaminergic (DA) transmission of the VTA in this regulation. Male Wistar rats (n=35) in urethane anaesthesia received an intra-VTA microinjection of either flupenthixol (FLU; doses of 5.0, 2.5, 1.25 and 0.625 µg) or amphetamine (AMPH; 2.5 and 5.0 µg) following control solvent microinjection. Peak power (Pmax) and corresponding peak frequency (Fmax) for delta and theta bands were extracted from EEG recording. Flupenthixol at a dose of 1.25 µg evoked long-lasting theta, continuing for 32.0 min on average, with a mean latency of 7.1 min. Other doses of FLU caused an increase of Pmax theta and reduction of Pmax delta without generating visually recognizable, regular theta rhythm. 5 µg of AMPH evoked theta continuing for 24.4 min on average, with a mean latency of 9.7 min. The lower dose was much less effective, with its outcome resembling the one after the less active FLU doses. During pharmacologically induced theta rhythm, both after FLU and AMPH, brief episodes of asynchronous activity appeared periodically, and they were more frequent and longer in AMPH groups. AMPH may act locally on multiple sites, inhibiting DA cells in somatodendritic region but also increasing dopamine release in target structures, and this, depending on AMPH dose, can lead to induction of theta rhythm. Locally administered DA antagonist on the other hand, when used at a proper dose, can produce theta most likely by the mechanism of inhibiting autoreceptors.

Induction of hippocampal theta rhythm by electrical stimulation of the ventral tegmental area and its loss after septum inactivation.

The ventral tegmental area (VTA), which may be one of the structures involved in regulation of hippocampal theta rhythm, sends direct projections to the hippocampus and also to the forebrain septum, the key centres involved in theta generation. In the present study we aimed at assessing which projections from the VTA (direct or through the septum) participate in regulation of hippocampal electric activity. Experiments were conducted on 3 groups of urethanised male Wistar rats. In the first group (n=6) electrical stimulation of the VTA was used to evoke theta rhythm episodes in hippocampus. Stimulation was repeatedly applied in control conditions and after procainic blockade of the septum. The second group (n=6), subjected to unilateral electrical stimulation of the VTA (30-s stimulation at 10-min intervals during 2h) and to subsequent detection of Fos expression, served to measure neuronal activation of the target mesolimbic structures. Activation levels of selected structures were compared to data from analogous stimulation of the zona incerta (ZI, the third group, n=6). Stimulation of the VTA immediately generated regular theta rhythm in both hippocampi. Inactivation of the septum with procaine temporarily abolished this effect. VTA stimulation increased the density of Fos in the ipsilateral nucleus accumbens. Stimulation of the ZI never generated theta but evoked significant induction of Fos expression in the hippocampus. Our data suggest that the projection through which the VTA enhances theta rhythm is not direct but is incorporated into the main route of theta generation, which involves septum as the main relay node.

Involvement of GABAergic transmission in the midbrain ventral tegmental area in the regulation of hippocampal theta rhythm.

Previously we indicated that the ventral tegmental area (VTA) may belong to the system regulating hippocampal theta rhythm. In the present study, we aimed at assessing the role of the GABAergic system of the VTA in regulation of hippocampal electric activity. Male Wistar rats received unilateral intra-VTA microinjection of either bicuculline (50ng/0.5µl, n=9), muscimol (100ng/0.5µl, n=10) or phaclofen (500ng/0.5µl, n=9). 1-min tail pinch stimulations were applied at 10-min intervals to evoke theta rhythm episodes in hippocampus. We analysed peak power (P(max)) and corresponding frequency (F(max)) of EEG signal at delta and theta bands. Bicuculline induced theta rhythm in both hippocampi with 0 latency, continuous for ca. 33min. Phaclofen also induced theta but in this group it appeared with latency (17.45±3.16min on average), lasted for ca. 33.6min and during this time was interrupted by periods of irregular activity of variable length. Tail pinch was not applied in these groups. Muscimol induced an opposite effect: depression of theta P(max) with simultaneous increase in delta P(max) and a decrease in F(max) delta during episodes of tail pinch-evoked theta. This effect had variable latency and no return to the control EEG could be observed. We propose that GABA activity in the VTA is of tonic character, so that abolition of this mechanism produces immediate effect, i.e. theta induction (strong by GABA(A) and weak by GABA(B) receptors blockade), whereas enhancing the already present GABAergic inhibition causes delayed, prolonged changes expressed as gradual loss of theta synchronisation.

Modulation of hippocampal theta rhythm by the opioid system of the pedunculopontine tegmental nucleus.

The pedunculopontine tegmental nucleus (PPN) belongs to the brainstem system which synchronizes hippocampal activity. Theta relevant intra-PPN circuitry involves its cholinergic, GABA-ergic and glutamatergic neurons and Substance P as neuromodulator. Evidence that PPN opioid elements also modulate the hippocampal theta is provided here. In urethane-anesthetized rats a unilateral microinjection of morphine (MF) (1.5 and 5 microg) increased the maximal peak power of tail pinch-induced theta. The higher dose also increased the corresponding frequency. When the theta was evoked by intra-PPN injection of carbachol (10 microg), the addition of MF (5 microg) prolonged theta latency and shortened the duration of the theta. These effects of MF were blocked by naloxone (5 microg). The results obtained suggest that the PPN opioid system can enhance or suppress the hippocampal theta depending on the actual level of PPN activation.