Re-afferent effects of individual static and dynamic γ-stimuli during maintained fusimotor stimulation

The ability of maintained dynamic and static fusimotor stimulation to modulate the primary afferent response of the muscle spindle in the rhythm of γ-stimulation was investigated using a highly sensitive method for modulation detection. The effect of 41 γ-fibers (13 dynamic; 28 static) on 38 primary afferents obtained from the tibialis anterior muscle of the cat was studied. It was found that maintained stimulation of 10 out of the 13 dynamic (77%) and of 25 out of the 28 static (89%) γ-fibers could evoke significant modulations of the primary afferent response in the rhythm of fusimotor stimulation at a minimum of one stimulation rate. Moreover, both static and dynamic γ-stimulations could evoke significant primary afferent modulations almost over the entire range of stimulation rates studied (30–300 stimuli per second). These results show that both γ-systems can modulate the primary afferent response in the rhythm of fusimotor stimulation over a wide range of stimulation rates; thus the central nervous system may be provided with re-afferent information about the effect of each individual γ-motoneuron discharge. Some hypotheses for the internal spindle mechanism responsible for the afferent modulations are discussed.     

Septo-hippocampal relationships during EEG theta rhythm

The firing pattern of 96 neuron in the medial septal nucleus (MSN) and diagonal band of Broca (DBB) and their functional relationships with hippocampal theta generators were investigated in rats during physostigmine-induced hippocampal theta rhythm (theta). Three types of discharge were found. (a) Type 1 (74%), in rhythmic bursts phase-locked with theta. (b) Type 2 (19%), non-rhythmic but phase-related to theta. (c) Type 3 (7%), unrelated to theta and non-rhythmic. Three sub-patterns of type 1 discharges were found. (1) Type 1A (56%), with regular bursts at high intraburst rates. (2) Type 1B (11%), with short bursts, or a single spike per theta cycle, and long interburst silent intervals. (3) Type 1C (6%), without clearly separate bursts, but with a rate modulation related to theta. Electrical stimulation of structures projecting to MSN-DBB or hippocampus tended to determine a constant poststimulus phase of theta and type 1 unit activity (i.e., reset). Our findings are in agreement with the hypothesis that MSN-DBB type 1 cells function as the theta "pacemaker', type 1A probably being the most influential in generating theta. Type 1B may be local interneurons and type 1C may receive rhythmic information from both septal and hippocampal neurons. Type 2 cells, although non-rhythmic, carry information of theta rhythmicity. Stimulus effects suggest that hippocampal theta reset is generated in the MSN-DBB.     

Cross-correlation analysis of septohippocampal neurons during theta-rhythm

The activity from 55 septohippocampal neuron pairs was examined in rats anesthetized with urethane. In addition to the statistical characterization of the firing patterns of the recorded units, the functional interactions between pairs of neurons and between neurons and hippocampal theta (theta) waves were investigated with cross-correlation techniques. Pairs were classified according to the rhythmic or non-rhythmic discharge pattern of their neurons. (a) theta-Pairs were those in which both the medial septal (MS) and hippocampal (HPC) units were rhythmic (type 1 units). (b) Pairs with a rhythmic and a theta-related non-rhythmic unit (type 2 unit) were called mixed pairs. (c) Pairs composed of type 2 units were called type 2 pairs. theta-Pairs showed periodic cross-correlations and frequently fired with a phase difference which could change in different pairs. Mixed pairs also showed periodic cross-correlations although one of the units was non-rhythmic. Type 2 pairs showed non-periodic positive cross-correlations. Our data provide new information regarding the temporal relationship between MS and HPC rhythmic activities supporting the role of the MS in providing the afferent timing for the generation of theta-rhythm in the HPC.     

Hippocampal EEG responses induced by carbachol and atropine infusions into the septum and the hippocampus in the urethane-anaesthetized rat

Infusion of 1 μg of carbachol, a potent cholinergic agonist, into the lateral septum of the urethane-anaesthetized rat systematically caused the induction of clear-cut hippocampal theta (θ). However, infusion of an equivalent amount of the drug into the hippocampus, close to the recording electrode, failed to induce θ in 50% of the animals and produced a mixture of θ waves and desynchronized activity, resulting in atypical EEG patterns, in the remaining subjects. Both carbachol EEG effects were blocked by intraseptal infusion of the antimuscarinic agent, atropine. Our data demontrate that muscarinic receptors in the septum are predominent sites for cholinergic agonist-antagonist action capable of generating or suppressing hippocampal θ in the rat. They also indicate that intraseptal cholinergic mechanisms play an important role in the initiation and generation of this rhythm.     

Influences of hypothalamic stimulation upon septal and hippocampal electrical activity in the cat.

Spontaneous patterns of hippocampal EEG and septal cell activity were studied in immobilized cats, and the influences of high frequency stimulation of medial hypothalamus (MH) and lateral hypothalamus (LH) were determined. Septal cells were divided into 3 classes on the basis of their discharge patterns: (1), rhythmic bursting (2), non-rhythmic bursting and (3), non-bursting, and the relationship of these discharge patterns to hippocampal theta rhythm was analyzed. Rhythmic bursting cells displayed close frequency and phase relations to hippocampal theta rhythm and were located chiefly in the diagonal band of Broca. Cells of the other two categories were found both within and outside of the diagonal band region.     

Electrophysiological analysis of synaptic transmission between intraocular hippocampus/locus coeruleus co-transplants

We have examined the establishment of functional connectivity between hippocampal and locus coeruleus co-transplants in oculo. Co-transplants were allowed to mature in oculo for 8–54 weeks following grafting and were subsequently removed from the anterior eye chamber for in vitro electrophysiological studies. Single hippocampal transplants in oculo have been shown to exhibit prolonged synaptic responses to local electrical stimulation¹⁷, and similar responses were observed in hippocampal neurons following stimulation of the hippocampal portion of hippocampus-locus coeruleus co-transplants. Electrical stimulation of the locus coeruleus attenuated the afterhyperpolarization in hippocampal neurons elicited by the injection of depolarizing current, an effect that has been described previously in hippocampal slices following direct application of norepinephrine, and this effect was antagonized by pretreatment with the β-adrenergic antagonist timolol. Stimulation of the locus coeruleus also produced both hyperpolarizing and depolarizing changes in the resting membrane potential in hippocampal neurons in 2- and 6-month-old co-transplants. In the 2-month-old co-transplants the responses were primarily depolarizing, and appeared to be mediated by a β-adrenergic receptor, whereas in the 6-month-old co-transplants the responses were more varied. The results suggest that functional α- as well as β-adrenergic receptors develop in oculo, and that the release of norepinephrine at nerve terminals in double grafts produces effects in the hippocampal neurons which are similar to those observed during superfusion of the hippocampal slice preparation with exogenous norepinephrine.     

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 μM carbachol on hippocampal slices. Fifty-six percent of the cells in this sample were termed theta (θ)-related, ie., they exhibited membrane potential oscillations of 5–28 mV and rhythmic spike discharges related to the carbachol-induced extracellular θ-rhythm. The remaining 44% of the cells did not show the above relationship to the extracellular θ-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 θ in hippocampal slices. This preparation will be a valuable model for studying cellular mechanisms and network properties underlying electroencephalographic activity.     

Synaptic contributions to θ rhythm genesis in rat CA1–CA3 hippocampal pyramidal neurons in vivo

The effects of intracellular Cl⁻ diffusion and hyperpolarizing current pulses on inhibitory postsynaptic potentials (IPSPs) and the transmembrane theta rhythm of CA1–CA3 pyramidal neurons were tested in urethanized and curarized rats. Cl⁻ diffusion and hyperpolarizing currents decreased the amplitude of IPSPs evoked by fornix stimulation without modifying the θ rhythm amplitude and phase. The membrane conductance was typically 22–46% higher at the positive than negative intracellular θ peaks. Results indicate that in curarized rats excitatory postsynaptic potentials were the main components of intracellular θ without an important participation of IPSPs in θ rhythm genesis.     

Cingulate cell discharge patterns related to hippocampal EEG and their modulation by muscarinic and nicotinic agents

Surface to depth recordings of slow wave θ activity were made through the posterior cingulate cortex (area 29). In other experiments the discharge patterns of neurons located throughout this region were recorded during the occurrence of large amplitude irregular activity (LIA) and slow wave θ activity (type 2) in the hippocampal formation of the urethane-anesthetized rat. The response of these neurons to the systemic administration of the cholinergic agonists, eserine and nicotine, was also tested. The majority of cells (80%) related to hippocampal EEG states were classified as tonic θ-on cells (non-rhythmic). These cells increased their discharge rates significantly, when the slow wave activity in the hippocampus changed from LIA to θ. The administration of eserine and nicotine induced slow wave θ in the hippocampus accompanied by the increased discharge rate of tonic θ-on cingulate cells. The excitatory action of nicotine on the discharges of tonic θ-on cingulate cortical cells is in direct contrast to its inhibitory action on phasic and tonic hippocampal θ-on cell discharges. The observation that phase reversals did not occur in area 29, together with the low incidence of phasic (rhythmic) θ-on cells, suggests that the posterior cingulate cortex does not independently generate type 2 θ. Possible significance of these findings for the interactive functioning of the hippocampal formation and posterior cingulate cortex is discussed.     

Ischemia-induced changes in α-tubulin and β-actin mRNA in the gerbil brain and effects of bifemelane hydrochloride

Using in situ hybridization histochemistry, we examined changes in the cytoskeletal protein α-tubulin and β-actin mRNAs in the gerbil brain 14 days after transient ischemia. In an attempt to identify the changes induced in the synthesis of cytoskeletal protein by schemia, we also evaluated the effects of post-ischemia administration of bifemelane on these cytoskeletal proteins. α-Tubulin and α-actin mRNAs were decreased in the CA1 region 14 days after transient ischemia. These decreases coincided with the loss of CA1 pyramidal cells, suggesting that they may have been related to delayed neuronal death. The β-actin mRNA level in ischemic controls was significantly increased in the dentate gyrus, habenular nucleus, and medial and lateral thalamic nuclei, where some afferent nerves project into the hippocampal pyramidal cells. The increased β-actin mRNA suggests that there may be a compensatory enhancement of actin synthesis in the afferent neurons that restores loosened synaptic connections with the ischemic cells in the CA1-4 fields. Administration of bifemelane just after recirculation prevented most of the ischemia-induced mRNA reductions in the CA1 field. Bifemelane's effect may be related to inhibition of Ca²⁺ influx and its radical scavenging activity. When bifemelane was administered to the ischemic group, α-tubulin mRNA levels significantly increased in the dentate gyrus and amygdaloid nucleus, and β-actin mRNAs showed a tendency to increase in the CA3 and CA4 fields, dentate gyrus, and medial and lateral thalamic nuclei. These findings suggest that bifemelane may enhance synthesis of cytoskeletal protein, especially in the ischemic brain, inducing axon outgrowth or synapse formation.     

Reset of hippocampal rhythmical activities by afferent stimulation

Physostigmine induced theta rhythm and unit activity were recorded from the dorsal hippocampus in immobilized locally anesthetized rats. Correlations between theta and rhythmical unit activity and their modifications by hippocampal afferent stimulation were studied. The principal finding was that electrical stimulation of afferents reset theta and rhythmical unit activity in phase. Poststimulus theta displayed a variable frequency which depended upon the structure stimulated. Lower frequencies were evoked by septal, higher frequencies by entorhinal and reticular formation stimulations. When theta rhythms were absent either by spontaneous disappearance or as a consequence of lesions in the fornix superior of septum, the reset was not observed. The reset of the theta rhythms and unit activity by afferent stimulation, suggests that the hippocampus may participate in timing mechanisms.     

Extracellular calcium and action potentials of soma and dendrites of hippocampal pyramidal cells

Calcium activity ([Ca2+]0) and focal potentials were recorded from dendritic and cell body layers of CA1 and CA3 regions of slices of hippocampal tissue. Responses were evoked by focal stimulation of afferent fiber bundles. In order to evoke a detectable decrease of [Ca2+]0, stimulation had to be intense enough to cause postsynaptic discharge of action potentials; evoking EPSPs alone was insufficient. Responses of [Ca2+] were consistently greater in stratum pyramidale than in stratum radiatum. It is concluded that in hippocampal tissue activation of soma and proximal dendritic membranes is the most important contribution to the decrease of [Ca2+]0 in response to afferent stimulation.     

Analgesic effect of intrathecal morphine demonstrated in ascending nociceptive activity in the rat spinal cord and ineffectiveness of caerulein and cholecystokinin octapeptide

The effect of intrathecal injections of morphine and the two peptides, caeruelin and cholecystokinin octapeptide (CCK-8), on the activity in ascending axons of the spinal cord evoked by electrical stimulation of primary nociceptive afferents was studied in spinal rats with decerebration. Morphine (20 μg) depressed the spontaneous activity and the activity evoked from either Aδ- or C-fibres. The co-activation by Aδ-fibre stimulation of ascending axons activated by stimulation of C-fibres and the activity in ascending axons activated by stimulation of afferent Aβ-fibres were not influenced by morphine. C-Fibre-evoked ascending activity was also depressed by morphine (10 μg and 5 μg). Ascending nociceptive activity was not changed by caerulein (30 ng) and CCK-8 300 ng, but it was depressed by a subsequent injection of morphine (20 μg). The depressant effects of morphine were abolished by an intravenous injection of naloxone which, when given alone, facilitated the ascending nociceptive activity. It is concluded that: (1) an intrathecal injection of morphine selectivity depressed the ascending nociceptive activity; (ii) the depression produced by morphine is an equivalent for spinal analgesia following intrathecal injection of morphine to man; and (iii) the two components of the spinal nociceptive system, the motor and the sensory path, can independently be influenced by drugs.     

“Epileptic” brain damage in rats induced by sustained electrical stimulation of the perforant path. I. Acute electrophysiological and light microscopic studies

Sustained electrical stimulation of the perforant path in urethane-anesthetized rats evoked hippocampal granule cell population spikes and epileptiform discharges. After stimulation, recurrent inhibition in the granule cell layer was abolished. Light microscopic analysis revealed a highly reproducible pattern of hippocampal damage to dentate pyramidal basket cells, hilar cells in general and CA3 and CA1 pyramidal cells. CA2 pyramidal cells and dentate granule cells were relatively unaffected. When perforant path stimulation on one side of the brain evoked bilateral granule cell discharges, damage was bilateral. Unilateral hippocampal seizures were associated with unilateral hippocampal damage. Rapid Golgi-stained hippocampi exhibited spherical dendritic swellings at the sites of termination of excitatory entorhinal afferents to the hippocampus and in the mossy fiber region. Electrical stimulation of a single excitatory afferent to the hippocampus appears to reproduce the “epileptic” pattern of hippocampal damage without using convulsant drugs and without causing motor convulsions. It is suggested that seizure-associated brain damage in caused by excessive pre-synaptic release of excitatory transmitter that induces intracellular post-synaptic changes that lead to dendritic swelling and cell death.     

Effect of stimulation in the periaqueductal grey matter on activity in ascending axons of the rat spinal cord: selective inhibition of activity evoked by afferent Aδ and C fibre stimulation and failure of naloxone to reduce inhibition

In rats with prenigral decerebration, the effect was studied of electrical stimulation of the periaqueductal grey matter (PAG) on the activity recorded from axons ascending in the spinal cord. These axons were activated by electrical stimulation of afferent Aβ, Aδ and C fibres in the ipsilateral sural nerve.Stimulation of the PAG with trains of impulses by itself evoked ascending activity, but strongly depressed the impulse transmission from C fibres to neurones with ascending axons. It exerted a weaker effect on impulse transmission from Aδ fibres and had no effect on impulse transmission from Aβ fibres to neurones with ascending axons. Intravenous naloxone, 1 mg/kg, did not diminish the depressant effect of PAG stimulation.Intravenous morphine depressed the activation of ascending axons from afferent C fibres (0.5 mg/kg) more markedly than that from afferent Aδ fibres (2 mg/kg), but did not modify the depression of ascending activity produced by PAG stimulation. Naloxone antagonized the depressant effect of morphine.The results indicate that PAG stimulation inhibits ascending activity evoked by noxious stimuli by a mechanism which does not necessarily involve endogenous opiates.     

Detection of an atropine-resistant component of the hippocampal theta rhythm in urethane-anesthetized rats

An important pharmacological feature of the hippocampal θ rhythm in urethane-anesthetized animals is its apparent sensitivity to antimuscarinic drugs. This sensitivity may be partly due to a masking of the θ frequency by increases in both higher and lower frequency EEG components that are unrelated to any residual θ rhythm. The discovery of atropine-resistant, rhythmic medial septal neurons has provided a physiological trigger for averaging EEG and unit activity after large atropine doses. Such averaging has permitted the detection of an atropine-resistant component of the hippocampal θ rhythm in urethane-anesthetized rats. The postatropine θ activity recorded from both CA1 (superficial to the pyramidal cell layer) and dentate (near the hippocampal fissure) in 15 rats was typically reduced in amplitude, but the recordings from the two locations maintained their phase relations to the septal units and to each other. The presence of this residual θ component after doses as large as 100 mg/kg indicates that it cannot be mediated by muscarinic cholinergic receptors. The coupling of the signal to the atropine-resistant septal cells strengthens our previous suggestion that these septo-hippocampal neurons are not cholinergic, and are therefore probably GABAergic.     

α7-Nicotinic receptor expression and the anatomical organization of hippocampal interneurons

C3H and DBA/2 mice differ in their hippocampal inhibitory function, as measured by the inhibitory gating of pyramidal neuron response to repeated auditory stimulation. This functional difference appears to be related to differences in expression of the α7 nicotinic cholinergic receptor, which may be generally expressed by interneurons. This study examines the relationship between genetic variation in α7 receptor subunit expression and GABAergic interneuron distribution in various regions and layers of the hippocampus in the two mouse strains. Subpopulations of hippocampal interneurons in both mouse strains were found to bind [¹²⁵I]α-bungarotoxin. However, the distribution of the [¹²⁵I]α-bungarotoxin-positive hippocampal interneurons was significantly different between C3H and DBA/2 mice. In region CA1, and to a lesser extent in region CA3, DBA/2 mice had increased numbers of [¹²⁵I]α-bungarotoxin-positive neurons in stratum lacunosum-moleculare and decreased numbers in stratum oriens. Similar differences in GABAergic neuron distribution were observed in region CA1 in the two strains. C3H/DBA/2 F1 animals were backcrossed to the C3H parental strain for six generations, with selection for either the DBA/2 or C3H allelic variant of the α7 receptor gene. The distribution of [¹²⁵I]α-bungarotoxin labeling closely resembled the DBA/2 parental phenotype in animals retaining the DBA/2 allele of the α7 gene. These data suggest that the α7 receptor gene locus may influence the anatomical organization of at least a subset of hippocampal interneurons by an as yet unidentified mechanism. This difference in interneuron anatomy may also contribute to functional differences in inhibitory sensory gating between the two strains.     

Enhancement of afferent fiber activity in hippocampal slices

The potentiation of synaptic activity in the hippocampal formation is a well documented phenomenon. It has been suggested, however, that a recruitment of additional afferent fibers can contribute to such an increase in synaptic activity. The hypothesis of an enhancement in afferent fiber activity, therefore, was investigated with the in vitro hippocampal slice preparation. Isolated radiatum fiber compound action potentials were evoked with paired electrical stimuli of equal intensity and separated in time by 30–40 msec. Statistical comparisons between control and test evoked responses reveal an augmentation of the test responses in support of the hypothesis of an enhancement in afferent fiber activity.