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Effects of early undernutrition on dendritic spines of cortical pyramidal cells in the rat.

Restricted food intake in male rats between birth and 20 days of age reduced the density of spines of layer V pyramidal cells in the frontal, parietal and occipital cortices. The loss of dendritic spines was more marked in pyramidal cell areas receiving projections from nonspecific multisynaptic systems than in cell areas associated with specific afferent systems or in areas receiving callosal projections. It was suggested that the differential effect of undernutrition in nonspecific, specific and callosal afferent projection areas of the pyramidal cortex might be related to the impairment of brain integrative functions reported to follow perinatal undernourishment.

Animals

Cerebral ischemia induces transient intracellular redistribution and intranuclear translocation of the raf proto-oncogene product in hippocampal pyramidal cells.

In this report we describe changes in the intracellular redistribution of raf serine/threonine protein kinase (product of the raf proto-oncogene family) in hippocampal neurons following cerebral ischemia in Mongolian gerbils. For immunohistochemical localization studies polyclonal antisera specific for each of the A, B, and Raf-1 isotypes of raf, as well as a pan-raf antisera, were employed. Of these, only sera recognizing B-raf, as well as the general v-raf (raised against the conserved C-terminal region) were positive, indicating that B-raf is the major isotype in this neuronal region. Three different ischemic models were used (repeated 3 times for two min and single 5 or 15 min occlusions, of the common carotid arteries) to demonstrate that ischemic insult causes redistribution of raf protein kinase into the cell nucleus of hippocampal neurons. Increased amounts of raf protein in the nuclei of pyramidal cells following ischemia was confirmed by Western blot analysis of isolated nuclear fractionations. Moreover, an elevation in the level of nuclear raf protein also was detected in the contralateral (i.e. non-occluded hemisphere) neurons of CA1 and CA3 subfields 4 days after the ischemic insult indicating a possible transsynaptic increase in the amount of raf protein along with redistribution. The intranuclear translocation of the immunoreactive material started from the perinucleolar rim and with time extended throughout the nucleus. Enhanced levels and altered redistribution of the raf polypeptide in the nuclei of pyramidal cells of the CA3 subfield appears to be reversible and returns to the normal level 12 days following the ischemic insult.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence

Increased glucose metabolism during long-duration recurrent inhibition of hippocampal pyramidal cells.

The locally subnormal brain metabolism observed in some experiments utilizing the Sokoloff 2-deoxyglucose (2-DG) method has often been attributed to postsynaptic inhibition despite the fact that inhibitory postsynaptic potentials are themselves caused by energy-requiring mechanisms. To explore this issue, neurophysiologically confirmed long-duration recurrent inhibition of hippocampal pyramidal unit firing was induced by low frequency (2 to 4 Hz) stimulation of the fornix for 60 min following intravenous infusion of [14C]-2-DG. The resulting autoradiograms showed that long-duration suppression of pyramidal cell firing was accompanied by distinctly increased hippocampal 2-DG uptake, particularly in the stratum pyramidale, which contains a dense plexus of inhibitory interneuronal terminals upon pyramidal cells. Both the pyramidal inhibition and the increased 2-DG uptake were confined to the ipsilateral hippocampus in animals with previously severed fornices and hippocampal commissures. In a second series of rats, the excitatory entorhinohippocampal "perforant path" (PP) was stimulated at low frequency (2 to 9 Hz) following 2-DG administration. At 2 to 4 Hz, each PP stimulation resulted in a brief burst of pyramidal unit firing followed by short-duration firing suppression; this result was associated with paradoxically decreased 2-DG uptake in the ipsilateral stratum molecular. By contrast, 7 to 9 Hz entorhinal stimulation induced PP-mediated excitation immediately followed by powerful intrinsic hippocampal inhibition, evidenced by prolonged pyramidal unit suppression after each stimulation. This suppression was accompanied by increased 2-DG uptake in the dentate stratum molecular and hippocampal stratum pyramidale. Thus it appeared that even with entorhinal stimulation, hippocampal 2-DG uptake was more closely associated with long-duration recurrent inhibition than with transient pyramidal excitation. Therefore, although it still remains possible that regions of hypometabolism observed in some previous 2-DG studies may actually reflect mild inhibition, other mechanisms such as disfacilitation are more likely mechanisms for this metabolic pattern.

Animals

The locomotor and exploratory activities in rats after lesion of hippocampal pyramidal cells with kainic acid.

The effect of single kainic acid (KA) intracerebroventricular(icv) injection on spontaneous locomotor, exploratory, basal and total activities in rats measured a different times was tested. Whereas at 3 hr after KA icv administration an increase of the total, basal and spontaneous activities of rats were observed, a great decrease of the exploratory activity was noted. KA at 24 hr after its administration depressed all types of rats activities. From 72nd hr after KA icv injection a significant increase in the total, exploratory and spontaneous locomotor activities without changes in the basal activity was observed. The above mentioned changes persisted up to 10-20 days after KA administration. The histological examinations showed evident neuropathologic changes in hippocampal stratum pyramidale. The most evident changes concerned the hippocampal areas CA3/CA4 and CA1. There were observed shrinking, narrowing and fragmentation of pyramidal cells. An intense progressive gliotic reaction was present among the fragmented cells, the layer of pyramidal cells being considerably diminished.

Animals

Visibility of synaptically induced conductance changes: theory and simulations of anatomically characterized cortical pyramidal cells.

A recent report has provided evidence that there are no significant increases in the neuronal input conductance during the response of cortical cells in cat visual cortex to non-preferred visual stimuli (Douglas et al., 1988). A criticism of experiments of this kind is that changes in the membrane conductance occurring in the dendritic tree may not be visible from electrodes that impale the soma. Our paper describes theoretical and numerical results concerning the visibility of synaptically induced conductance changes from intracellular electrodes, in both ideal and anatomically well-characterized cortical neurons. Based on earlier work by Rall (1967), we here derive theoretical expressions for the change in input conductance at any location in a passive dendritic tree resulting from activation of a single synapse and obtain bounds for the effects of multiple synapses. We find that the conductance change measured at the cell body is always less than the sum of the synaptic conductance changes and that this observed conductance change does not depend on the synaptic reversal potential. For the case of an infinite dendritic cylinder, the change in input resistance due to a single synaptic input decays exponentially with distance of the synapse from the recording site. Numerical simulations of synaptic inputs that change approximately as fast as the membrane time-constant produce an increase in input conductance that is only slightly less visible than that of a constant input. We also compute the changes in somatic input conductance of 2 morphologically identified pyramidal cells from cat visual cortex during activity of a single inhibitory basket cell with known synaptic input locations. We find that the increase in conductance due to the activity of the inhibitory basket cells is clearly visible from the cell body of the pyramidal cells and that a 70% reduction in the amplitude of excitation is associated with at least a 30% increase in somatic input conductance, which would be visible in intracellular recordings. Taken together with the negative experimental evidence of Douglas et al. (1988), our results cast doubt on a large class of models of direction selectivity that rely on synaptically mediated inhibitory conductance increases to veto or block excitatory conductances increases.

Animals

Morphological changes of the pyramidal cell nucleolus and nucleus in hamster frontal cortex during development and aging.

The development and aging of the nucleolus and nucleus in layer V pyramidal cells in the hamster cerebrum were studied by light and electron microscopy. The nucleoli appeared in the newborn as occasional fibrillar masses adjacent to peripherally placed bodies of chromatin. By maturity, a single, generally central, nucleolus proper with nucleolus-associated chromatin was present. Nucleolar microbodies were observed at 10, 15, 20 and 480 days, but not in the newborn, 5-or 90-day animal. An intranucleolar body was not observed at the electron at the electron-microscopy level in these pyramidal cell nucleoli at any age in this series, in contrast to the situation in large motor neurons of the facial nucleus. The nucleus progressed from an irregular shape at birth to an oval shape at maturity. At 10 days, incipient invaginations of the nuclear membrane appeared; these subsequently increased in depth and frequency in the adult. The above changes, particularly in the nucleoli, are correlated in time with changes involving the endoplasmic reticulum. The correlations may indicate different periods of metabolic activity in the hamster pyramidal neurons. Four such periods can be differentiated on the basis of cytomorphic changes which may be correlated to reported development of function. The sequence of these changes, peculiar to the developing and aging hamster pyramidal neuron, differs from that seen in large spinal and cranial motor neurons. It appears that some features of nuclear immaturity, which are lost in larger neuronal types, are retained in the adult pyramidal neuron.

Aging

Morphologic abnormalities in the postnatal differentiation of CA1 pyramidal cells and granule cells in the hippocampal formation of the ataxic mouse.

The postnatal differentiation of the hippocampal formation of the ataxic mouse was studied. Brains from ataxic mice (axJ/axJ) and littermate controls (+/?), from 19 to 51 days of age, were either impregnated according to a Golgi-Cox procedure or sectioned and stained with Weil-hematoxylin and Darrow Red. The hippocampus and dentate gyrus, although somewhat reduced in cross-sectional area in the ataxic brain, appeared to have a normal complement of both pyramidal cells and granule cells, respectively. Examination of Golgi-Cox material showed significant differences in the differentiation of the dendritic tree of both pyramidal and granule cells. At 41 days the height of the apical dendrite of CA1 pyramidal cells in the ataxic brain was 76% of the control value, and the width was only 35%. Similarly, the basal dendritic tree was narrower in the ataxic mouse. In the dentate gyrus of the 41-day ataxic brain, the height of the granule cell dendritic tree was only 74% of the control value. These and other alterations in the dendritic morphology of both CA1 pyramidal cells and granule cells can be explained by a lack of growth of the dendritic tree during the developmental period studied. These findings are discussed in relation to other studies on intrinsic and extrinsic factors and their effect on normal hippocampal differentiation.

Animals

The effect of muscimol on hippocampal pyramidal cells.

The effects of muscimol on rabbit hippocampal pyramidal cell firing were studied and compared after iontophoretic, topical, and intravenous administration of the drug. All modes of application resulted in a bicuculline-sensitive, strychnine-insensitive, depression of the monosynaptically activated population spike evoked by micro-stimulation of the contralateral hippocampal field. These findings indicate that systemically administered muscimol selectively activates hippocampal GABA receptors suggesting that this compound may be useful for studying limbic system physiology.

Action Potentials

Axonal ramifications of hippocampal Ca1 pyramidal cells.

Intracellular injections of Lucifer Yellow into CA1 pyramidal cells of the in vitro guinea pig hippocampal slice enabled us to examine in detail the morphology of the axons of these neurons. We also recorded the electrophysiological responses of these neurons to alvear stimulation. In our morphological examinations, we found that many axons bifurcate in the alveus, with the major branch projecting caudally toward the subiculum and the second, thinner branch projecting rostrally toward the fimbria. Either axons may bifurcate further to produce several axon branches which follow parallel paths in the alveus. These axons also have local collaterals which project into strata oriens and pyramidale. In addition, a very fine plexus of axonal processes was observed in stratum oriens located largely within the basal dendritic field of the parent cell. Our electrophysiological experiments demonstrated that neurons could be activated antidromically by stimulation of the alveus at sites both rostral ad caudal to the neuron. Weak alvear stimulation occasionally evoked small potentials which appeared similar to fast prepotentials. The local axonal ramifications may be involved in recurrent pathways mediating feedback inhibition and/or excitation. The axonal bifurcations also may provide a basis for understanding the origins of fast prepotentials elicited with antidromic stimulation.

Action Potentials

Glutamate-induced action potentials are preceded by regenerative prepotentials in rat hippocampal pyramidal cells in vitro.

(1) The responses of CA1 pyramidal cells to short glutamate pulses (10-50 ms) delivered at sensitive spots in the apical dendrites have been analysed by intracellular recording. (2) The glutamate pulses elicited stable depolarizing responses in a dose- and frequency-dependent manner. (3) When a single action potential with a firing probability around 0.5 was elicited, a subtraction procedure showed that a slow depolarizing ramp preceded each spike. We call this ramp the glutamate-induced prepotential (GluPP). (4) In contrast to the upward convex subthreshold depolarization the GluPP was upward concave. (5) The GluPP amplitude and time course increased with depolarization of the membrane, a phenomenon which appears to be connected to the elevation of action potential threshold. (6) The GluPP was regenerative since once started, it ended in an action potential. (7) A specific N-methyl-D-aspartate receptor antagonist, DL-2-amino-5-phosphonovaleric acid (50 microM) reduced the glutamate-induced depolarization, but did not affect the form or amplitude of GluPP, once the latter was induced. (8) It is concluded that short glutamate pulses elicited action potentials through a prepotential mechanism, similar to the slow prepotentials induced by long depolarizing current pulses across the soma membrane. (9) A possible physiological role for the GluPP is discussed.

2-Amino-5-phosphonovalerate

Development of lamellar bodies and subsurface cisterns in pyramidal cells and neuroblasts of hamster cerebral cortex.

In pyramidal cells of hamster frontal cortex lamellar bodies and subsurface cisterns sequentially occurred during development from newborn to three months of age. Neither of these two specializations of the rough endoplasmic reticulum was seen in neurons of the newborn. The first specialization that we observed was the subsurface cistern, which appeared at five days and showed a significant increase both in frequency and in length throughout development. The lamellar body was first seen at ten days of age. This specialization showed a peaking of frequency, length, and number of cisterns per body at 15 days, which subsequently decreased gradually to 3-month-old levels. The occurrence of lamellar body-subsurface cistern complexes increased with age. We suggest that the lamellar body may be an ER specialization that is involved in an increased and/or stage-specific protein synthesis in the young prior to the final maturation of the usual neuronal protein synthetic organelles, and that in the 3-month-old neurons, the lamellar body may be involved in modifying, storing or transporting metabolites received from the neuropil components via the subsurface cisterns. The subsurface cistern, on the other hand, by virtue of its location subjacent to the neuronal plasma membrane and of its increased frequency from birth to maturation, may be involved in the exchange of metabolites and nonsynaptic forms of communication at all ages.

Aging

Dopamine action on hippocampal pyramidal cells.

Dopamine (DA) was applied to CA1 region pyramidal cells in slices of guinea pig hippocampus maintained in vitro in order to examine its electrophysiological effect on CNS neurons. DA induced hyperpolarization of membrane potential and an increased conductance in 75% the 21 CA1 neurons to which it was applied. DA also augmented the afterhyperpolarizations and increased conductance which normally follow spike trains in these neurons. These effects were not altered by intracellular injections of Cl- but were blocked when slices were bathed in Mn2+ solutions. The Mn2+ blockade of DA-induced hyperpolarizations could be overcome when large amounts of agonists were applied. The DA effects were long lasting, were mimicked by the dopamine agonists apomorphine and Epinine, and were blocked by the dopamine antagonists flupenthixol and chlorpromazine. Extracellular or intracellular application of cyclic AMP mimicked the effects of DA. The results suggest that DA-induced hyperpolarization and conductance changes are mediated by a Ca2+-activated K+ conductance. DA may increase the intracellular Ca2+ concentration through effects on one of the Ca2+ buffering mechanisms. The long duration of these effects suggest that DA works though some intracellular intermediary, perhaps cyclic AMP, considering that the actions of cyclic AMP on membrane properties are similar to those of DA. The dopaminergic projection to the hippocampus should have a powerful inhibitory action, which would be most effective in modulating the activities of neurons exhibiting high levels of excitability, particularly cells involved in cyclical burst generation.

Action Potentials

A new type of specific interneuron in the monkey hippocampus forming synapses exclusively with the axon initial segments of pyramidal cells.

By means of Golgi staining and gold-toning, we have found an interneuron in the pyramidal cell layer of the hippocampus which forms synapses exclusively on the axon initial segments of pyramidal neurons. An individual initial segment receives up to 30 symmetrical synapses from one axo-axonic cell. Each axo-axonic cell is in synaptic contact with the axon initial segments of several hundred pyramidal neurons. The interneuron is thus ideally situated to synchronize the output of a large population of pyramidal cells and so might be involved in the generation of rhythmic activity and in epileptogenesis.

Animals

The excitatory response of in vitro hippocampal pyramidal cells to normorphine and methionine-enkephalin may be mediated by different receptor populations.

As shown previously, opiate agonists increase the excitability of hippocampal pyramidal cells in a naloxone-reversible manner. In the present study, the degree of excitability was measured by population spike size recorded from hippocampus slices (CA1) obtained from naive or chronically morphinized rats. Cross tolerance could not be demonstrated to occur between met-enkephalin and normorphine in hippocampal cells made tolerant to morphine: the potent stimulatory effect of met-enkephalin remains when applied to hippocampal slices removed from chronically morphinized rats, whereas normorphine was no longer effective. When these slices are washed or exposed to naloxone a diminution of the population spike occurs. These results suggest that while both opiate agonists increase neuronal excitability of hippocampal pyramidal cells, this effect is most likely mediated via different receptor populations.

Animals

The non-pyramidal cells in layer III of cat primary auditory cortex (AI).

The form and location of non-pyramidal neurons in layer III of the primary auditory cortex (AI) of adult cats is described in Golgi, Nissl, and other material. The cells were compared to the profiles of retrogradely labeled, commissurally interconnected cells. A principal finding is that certain non-pyramidal and pyramidal cells project interhemispherically to AI; a second conclusion is that the retrogradely labeled commissural cells form small clusters or narrow strips separated by unlabeled patches even after massive injections in the opposite AI. The non-pyramidal cells of origin have not yet been conclusively identified, but they must include one (or more) of the following six types of cells observed in Golgi-impregnated material: tufted or bitufted cells with a radially elongated dendritic arbor; sparsely spinous stellate neurons with thin, smooth dendrites and vertically disposed axonal branches; small stellate cells with varicose dendrites, a restricted dendritic field, and a profusely branched local axon; bipolar neurons with long, thin dendrites; medium-sized multipolar cells with radiating, sparsely branched dendrites; and small stellate neurons with smooth dendrites and a tiny dendritic field. These non-pyramidal cells are found throughout layer III but are more numerous in the upper part, layer IIIa, where they mingle with the small pyramidal neurons. As a rule the axonal branches of non-pyramidal cells are more numerous than those arising from layer III pyramidal neurons, and although they have many axonal collaterals, most project locally and vertically in narrow radial strips. In contrast, pyramidal cell axons have ascending and descending components which invade large, lateral territories in many cortical layers. Layer III non-pyramidal neurons are similar to those in layer IV in certain respects, although their dendritic fields are more spherical and less tufted than those of layer IV cells, and their axons have more local, limited targets. These axons appear to contribute but little to the conspicuous, lateral fiber striae in layer III. The primary intrinsic targets of non-pyramidal cell axons appear to be the apical dendrites of medium-sized and large layer III pyramidal cells, and recurrent branches to the parent cell; their fine, distal branches fortify the vertical plexus in layer III, and certain axons may descend into layer IV. Since layer III in AI receives both commissural and thalamic input, it is possible that these parallel, afferent channels are to some degree segregated, and to some degree convergent, onto particular types of cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent Pathways

Morphine and opioid peptides reduce inhibitory synaptic potentials in hippocampal pyramidal cells in vitro without alteration of membrane potential.

We used intracellular recording in the hippocampal slice in vitro to characterize further the mechanisms behind the unusual excitatory action of opiates and opioid peptides on hippocampal pyramidal cells in vivo. No significant effect on resting membrane potential, input resistance, or action potential size in cortical area 1 (CA1) pyramidal cells was observed with morphine sulfate, beta-endorphin, [Met5]enkephalin, or [D-Ala2, D-Leu5]enkephalin at 1-50 microM. However, in all cells studied, these agents markedly reduced the size of inhibitory postsynaptic potentials generated by stimulation of the stratum radiatum or alveus. Excitatory postsynaptic potentials were also diminished in many of these cells. The effects of the opioids were antagonized by naloxone. These results are consistent with excitation of pyramidal neurons by a disinhibitory mechanism.

Animals

Synaptic sequences in mouse SmI cortex involving pyramidal cells labeled by retrograde filling with horseradish peroxidase.

A method combining anterograde degeneration and the retrograde transport of horseradish peroxidase (HRP) has been used to study synapses involving pyramidal cells in mouse SmI cortex. Neurons labeled with HRP are so well filled that even their finer processes, such as dendritic spines and axon collaterals, are clearly visible with both the light and electron microscopes. Results indicate that pyramidal cells projecting from SmI to ipsilateral MsI cortex receive thalamocortical input and have local axon collaterals which form asymmetrical synapses with spines and with varicose, non-spiny dendrites.

Animals

Effects of GABA on CA3 pyramidal cell dendrites in rabbit hippocampal slices.

Using the in vitro rabbit hippocampal slice preparation, we have investigated the effects of gamma-aminobutyric acid (GABA) iontophoresis on CA3 pyramidal cell dendrites. The predominant response (70% of the cells tested) was a hyperpolarization associated with a 30% decrease in cell input resistance (Rm). These hyperpolarizations displayed a very pronounced voltage dependency: they were decreased by cell depolarization and flattened by hyperpolarization. Bicuculline methiodide (BMI, 50 microM) did not abolish this response, nor did intracellular iontophoresis of chloride ions. In 5% of the cells, an additional hyperpolarization was obtained with longer ejection times; it reversed close to the reversal potential of the early component of the IPSP. In 25% of the cells, dendritic GABA application produced a depolarization. This response was reversed with cell membrane depolarization and was associated with a large (80%) decrease in Rm. The depolarizations were abolished by BMI (50 microM) and greatly increased by increasing the intracellular chloride concentration. None of the responses to GABA were affected by blockade of synaptic transmission. We conclude that the predominant response of CA3 pyramidal cell dendrites to GABA application is a hyperpolarization mediated by GABAB receptors and probably carried by potassium ions. The depolarizing responses are mediated via GABAA receptors and depend on an increase in chloride permeability.

Action Potentials