Fine structure and synaptic connections of identified neurons in the rat fascia dentata

Summary A survey is given of the synaptic connections of identified neurons in the rat fascia dentata based on our own Golgi/electron microscopic and light and electron microscopic immunocytochemical findings as well as on results obtained from the literature. The report largely deals with the dominating cell type in the region, the dentate granule cell. Of the various types of hilar cells, the GA-BAergic neurons, particularly the inhibitory basket cells, are taken into account. Differences in fine structure between granule cells and basket cells as well as mutual synaptic connections between these two types of dentate neurons are elaborated. This survey may provide a basis for further neurophysiological and pharmacological studies on these cells.     

Changing synaptic connections on cell bodies of growing identified spinal motoneurons of the eel, Anguilla

As the target musculature they innervate grows throughout life, certain segmental motoneurons from the spinal cord of Anguilla, readily identified on the basis of their form and position, also increase in size. In doing so, they present a steadily increasing target to the spinal and supraspinal neurons that innervate them. How the afferent neurons respond was assessed by measuring features of their synaptic boutons contacting the motoneuronal perikarya, as seen with electron microscopy. About 60% of the perimeter of the perikaryal profile of each motoneuron was found to be covered with synaptic bouton profiles, a value that is independent of the size of the motoneuron. Furthermore, the distances between synaptic profiles, their contact sizes (measured as apposition length) and the number and size of the vesicles each profile contains were all found to be relatively constant and also independent of motoneuronal size. In contrast, the number of synaptic profiles contacting a motoneuron correlated well with its perikaryal size. Our findings indicate that the challenge of a growing neuronal target is met by a steady increase in the number of contacting boutons, the form and spacing of which are held relatively constant; this strategy will require continual synaptic realignment at the target. Accepted: 18 September 2000     

Development of plastic mechanisms related to learning at identified chemical synaptic connections in Aplysia

In the marine snail Aplysia californica learned changes in behavior have been traced to alterations in synaptic efficacy. With the ability to raise the animals in the laboratory, we have explored the development of four types of plastic mechanisms at identified synapses: post-tetanic potentiation and pre-synaptic inhibition, which do not as yet have known behavioral functions, and homosynaptic depression, the cellular mechanism of short-term habituation, and heterosynaptic facilitation, the basis of short-term sensitization. Homosynaptic depression and pre-synaptic inhibition are present early in juvenile life. In contrast, post-tetanic potentiation and heterosynaptic facilitation appear only later, after a discrete interval. The step-wise ontogeny of synaptic plastic mechanisms in Aplysia parallels the gradual emergence of behavior in successive developmental stages. Interestingly, senescence reverses the developmental sequence for habituation and sensitization mechanisms. To pursue further an understanding of the relationship between synapse formation and plasticity underlying learning it will be necessary to extend these studies to dissociated cell culture where mechanisms can be explored on the molecular level. Cell culture may also permit examination of the development of cellular mechanisms underlying classical and operant conditioning which may clarify differences between associative and non-associative mechanisms.     

Role of endogenous cannabinoids in synaptic signaling

Research of cannabinoid actions was boosted in the 1990s by remarkable discoveries including identification of endogenous compounds with cannabimimetic activity (endocannabinoids) and the cloning of their molecular targets, the CB1 and CB2 receptors. Although the existence of an endogenous cannabinoid signaling system has been established for a decade, its physiological roles have just begun to unfold. In addition, the behavioral effects of exogenous cannabinoids such as delta-9-tetrahydrocannabinol, the major active compound of hashish and marijuana, await explanation at the cellular and network levels. Recent physiological, pharmacological, and high-resolution anatomical studies provided evidence that the major physiological effect of cannabinoids is the regulation of neurotransmitter release via activation of presynaptic CB1 receptors located on distinct types of axon terminals throughout the brain. Subsequent discoveries shed light on the functional consequences of this localization by demonstrating the involvement of endocannabinoids in retrograde signaling at GABAergic and glutamatergic synapses. In this review, we aim to synthesize recent progress in our understanding of the physiological roles of endocannabinoids in the brain. First, the synthetic pathways of endocannabinoids are discussed, along with the putative mechanisms of their release, uptake, and degradation. The fine-grain anatomical distribution of the neuronal cannabinoid receptor CB1 is described in most brain areas, emphasizing its general presynaptic localization and role in controlling neurotransmitter release. Finally, the possible functions of endocannabinoids as retrograde synaptic signal molecules are discussed in relation to synaptic plasticity and network activity patterns.     

Forskolin调节DHA对APP/PS1转基因小鼠突触可塑性和认知功能的增强作用

目的研究Forskolin调节DHA对APP/PS1转基因小鼠突触可塑性和认知功能的影响。方法对7月龄APP/PS1小鼠给予DHA及DHA和forskolin联合进行治疗。30d后,通过行为学试验检测其认知功能;通过组织病理学试验检测脑内细胞密度及脑突触形成能力和突触标记物Synaptophysin的表达以及通过Western blotting检测脑内pCREB1蛋白含量。结果 8月龄APP/PS1小鼠海马区面积、细胞密度、Synaptophysin和脑内pCREB1蛋白均明显降低,行为学上出现了明显的认知功能障碍。单独给予DHA以及DHA和forskolin联合给药治疗后,APP/PS1小鼠海马区面积、细胞密度、Synaptophysin和脑内pCREB1蛋白均增加,认知功能障碍得到改善,并且联合给药组的效果更为显著,但是均未恢复至生理水平。结论 DHA和forskolin联合治疗后,forskolin增强DHA对APP/PS1转基因小鼠突触病理和认知障碍的治疗作用。     

Synaptic connections of dentate granule cells and hilar neurons: results of paired intracellular recordings and intracellular horseradish peroxidase injections

Simultaneous intracellular recordings were made in the dentate gyrus of rat hippocampal slices, from pairs of the following cell types: granule cells, interneurons located in the granule cell layer, hilar interneurons, and spiny hilar "mossy cells". Granule cells were found to have strong excitatory effects on mossy cells and interneurons. Interneurons inhibited granule cells as well as other interneurons. No synaptic connections from mossy cells onto other cell types were found, within the confines of the slice, using intracellular recording methods. However, at the ultrastructural level, axon terminals of horseradish peroxidase-filled mossy cells were found making synaptic contacts in the hilus on dendrites of interneurons. These studies provide the first step towards determining the functional interactions of the various cell types in the fascia dentata.     

Reduction of neophobia in mice following lesions of the caudate-putamen

Electrolytic lesions of the caudate-putamen result in a significant decrease in neophobic responses in mice towards a novel object introduced into their familiar environment; however, preference for a novel environment was not altered by the lesion. These data provide a parallel between the effects of lesions of the caudate-putamen and the well-known "amygdala-lesion-syndrome." It is suggested that the striatal complex, which receives massive afferent projection systems, plays a crucial role in sensory-motor integration processes which allow the animals to adapt their responses towards biological significant stimuli in order to cope with their environment.     

Impaired synaptic plasticity and learning in aged amyloid precursor protein transgenic mice

We investigated synaptic communication and plasticity in hippocampal slices from mice overexpressing mutated 695-amino-acid human amyloid precursor protein (APP695SWE), which show behavioral and histopathological abnormalities simulating Alzheimer's disease. Although aged APP transgenic mice exhibit normal fast synaptic transmission and short term plasticity, they are severely impaired in in-vitro and in-vivo long-term potentiation (LTP) in both the CA1 and dentate gyrus regions of the hippocampus. The LTP deficit was correlated with impaired performance in a spatial working memory task in aged transgenics. These deficits are accompanied by minimal or no loss of presynaptic or postsynaptic elementary structural elements in the hippocampus, suggesting that impairments in functional synaptic plasticity may underlie some of the cognitive deficits in these mice and, possibly, in Alzheimer's patients.     

Sets of synaptic currents paired by common presynaptic or postsynaptic neurons

1. In the buccal ganglia of Aplysia, presynaptic neurons B4 and B5 produce similar inhibitory postsynaptic currents (PSCs) in several postsynaptic follower cells. Previous work has shown that both duration and amplitude of these PSCs vary, that each parameter may be altered transiently by manipulating presynaptic activity, and that these variations affect synaptic efficacy. 2. To permit synapse-to-synapse comparisons, the mean and coefficient of variation (CV) of both peak conductance (gpeak) and time constant of decay (tau) were determined for sets of synaptic currents evoked by direct intracellular stimulation of presynaptic neurons. For 56 synapses, gpeak = 0.40 +/- 0.33 (SD) microS for a CV of 0.83, and tau = 19.7 +/- 4.0 ms for a CV of 0.20. The synapse-to-synapse variability was within 5% of values obtained from a previous population. 3. The relative contributions of presynaptic and postsynaptic factors to efficacy and variability of PSCs were examined by recordings from two classes of three-cell networks and by comparing values of gpeak and tau at synapses sharing a common presynaptic or postsynaptic neuron. 4. In the first case, paired presynaptic inputs from B4 and B5 converged on a common postsynaptic cell. For 16 examples of this case, mean values of both gpeak and tau recorded in a single follower cell, but produced by different presynaptic neurons, were significantly closer than those recorded in different followers (P less than 0.001). The common postsynaptic cell did not constrain variability of these parameters; CVs for paired synapses were not significantly different from the population (P greater than 0.1).(ABSTRACT TRUNCATED AT 250 WORDS)     

Engagement of rat striatal neurons by cortical epileptiform activity investigated with paired recordings

The striatum is thought to play an important role in the spreading of epilepsy from cortical areas to deeper brain structures, but this issue has not been addressed with intracellular techniques. Paired recordings were used to assess the impact of cortical epileptiform activity on striatal neurons in brain slices. Bath-application of 4-amynopyridine (100 microM) and bicuculline (20 microM) induced synchronized bursts in all pairs of cortical neurons (< or = 5 mm apart) in coronal, sagittal, and oblique slices (which preserve connections from the medial agranular cortex to the striatum). Under these conditions, striatal medium spiny neurons (MSs) displayed a strong increased spontaneous glutamatergic activity. This activity was not correlated to the cortical bursts and was asynchronous in pairs of MSs. Sporadic, large-amplitude synchronous depolarizations also occurred in MSs. These events were simultaneously detected in glial cells, suggesting that they were accompanied by considerable increases in extracellular potassium. In oblique slices, cortically driven bursts were also observed in MSs. These events were synchronized to cortical epileptiform bursts, depended on non-N-methyl-D-aspartate (NMDA) glutamate receptors, and persisted in the cortex, but not in the striatum, after disconnection of the two structures. During these bursts, MS membrane potential shifted to a depolarized value (59 +/- 4 mV) on which an irregular waveform, occasionally eliciting spikes, was superimposed. Thus synchronous activation of a limited set of corticostriatal afferents can powerfully control MSs. Cholinergic interneurons located < 120 microm from simultaneously recorded MSs, did not display cortically driven bursts, suggesting that these cells are much less easily engaged by cortical epileptiform activity.     

The effects of cannabinoids on the brain.

Cannabinoids have a long history of consumption for recreational and medical reasons. The primary active constituent of the hemp plant Cannabis sativa is delta9-tetrahydrocannabinol (delta9-THC). In humans, psychoactive cannabinoids produce euphoria, enhancement of sensory perception, tachycardia, antinociception, difficulties in concentration and impairment of memory. The cognitive deficiencies seem to persist after withdrawal. The toxicity of marijuana has been underestimated for a long time, since recent findings revealed delta9-THC-induced cell death with shrinkage of neurons and DNA fragmentation in the hippocampus. The acute effects of cannabinoids as well as the development of tolerance are mediated by G protein-coupled cannabinoid receptors. The CB1 receptor and its splice variant CB1A, are found predominantly in the brain with highest densities in the hippocampus, cerebellum and striatum. The CB2 receptor is found predominantly in the spleen and in haemopoietic cells and has only 44% overall nucleotide sequence identity with the CB1 receptor. The existence of this receptor provided the molecular basis for the immunosuppressive actions of marijuana. The CB1 receptor mediates inhibition of adenylate cyclase, inhibition of N- and P/Q-type calcium channels, stimulation of potassium channels, and activation of mitogen-activated protein kinase. The CB2 receptor mediates inhibition of adenylate cyclase and activation of mitogen-activated protein kinase. The discovery of endogenous cannabinoid receptor ligands, anandamide (N-arachidonylethanolamine) and 2-arachidonylglycerol made the notion of a central cannabinoid neuromodulatory system plausible. Anandamide is released from neurons upon depolarization through a mechanism that requires calcium-dependent cleavage from a phospholipid precursor in neuronal membranes. The release of anandamide is followed by rapid uptake into the plasma and hydrolysis by fatty-acid amidohydrolase. The psychoactive cannabinoids increase the activity of dopaminergic neurons in the ventral tegmental area-mesolimbic pathway. Since these dopaminergic circuits are known to play a pivotal role in mediating the reinforcing (rewarding) effects of the most drugs of abuse, the enhanced dopaminergic drive elicited by the cannabinoids is thought to underlie the reinforcing and abuse properties of marijuana. Thus, cannabinoids share a final common neuronal action with other major drugs of abuse such as morphine, ethanol and nicotine in producing facilitation of the mesolimbic dopamine system.     

Lack of cross-tolerance to the antinociceptive effects of systemic and topical cannabinoids in morphine-tolerant mice

Opioids and cannabinoids produce antinociception through activity at spinal, supraspinal and peripheral sites. Tolerance to the antinociceptive effects of both the opioids and the cannabinoids develop when these agents are administered chronically. Although mutual potentiation of antinociceptive effects have been reported between opioids and cannabinoids, the development of antinociceptive cross-tolerance between these systems has not been demonstrated consistently. In the present investigation, we explored the possibility of antinociceptive cross-tolerance between systemic or topical morphine and systemic or topical cannabinoids in mice. Mice were made tolerant to morphine either by the subcutaneous (s.c.) implantation of a morphine pellet or repeated topical administration and then challenged with the mixed CB(1) and CB(2) receptor agonist WIN 55, 212-2 given s.c. or topically. Antinociception was indicated by increased tail-flick latencies to noxious radiant heat. Implantation with morphine pellets did not attenuate the antinociceptive potency of systemic or topical WIN 55,212-2. Moreover, twice-daily topical administration of morphine did not attenuate the antinociceptive potency of WIN 55,212-2 applied topically. These observations suggest that opioids and cannabinoids produce antinociception through mechanisms that are independent of each other at either the systemic or peripheral levels.     

Analysis of synaptic transmission at single identified boutons on rat spinal neurons in culture

The spatial organization of receptor channels has a major influence on the speed and possible plasticity of synaptic signal transmission. We have studied glutamatergic synapses on neurons in organotypic cultures of rat spinal cord. In order to avoid the problems related to the analysis of currents of unknown origin within a neuron, we chose to examine the functional properties of single identified synapses. Iontophoretic mapping of the cell surface revealed hot spots of high glutamate sensitivity coincident with presynaptic boutons stained with the dye FM 1–43. Local application of KCl to these sites caused bursts of synaptic release. Hot spots typically consisted of 330 receptors with an average single-channel conductance of 8.3 pS. Evoked synaptic currents involved only about 40–50 receptors and nevertheless showed characteristics of saturation. This suggests that glutamate receptor clusters at sites of presynaptic terminals are organized into well separated subclusters opposite release sites.This award-winning article is published as received and has not been subjected to the normal peer review process     

Presynaptic modulation of synaptic transmission by pregnenolone sulfate as studied by optical recordings

The effects of pregnenolone sulfate (PREGS), a putative neurosteroid, on the transmission of perforant path-granule cell synapses were investigated with an optical recording technique in rat hippocampal slices stained with voltage-sensitive dyes. Application of PREGS to the bath solution resulted in an acute augmentation of EPSP in a dose-dependent manner. The PREGS effect was dependent on the extracellular Ca(2+) concentration ([Ca(2+)](o)), but independent of NMDA receptor activation. PREGS caused a decrease in paired-pulse facilitation, which implies that PREGS positively modulates presynaptic neurotransmitter releases. Firmer support for this mechanism was that PREGS augmented the synaptically induced glial depolarization (SIGD) that reflects the activity of electrogenic glutamate transporters in glial cells during the uptake of released glutamate. The selective alpha7nAChR antagonist alpha-BGT or MLA prevented the SIGD increase by PREGS. Furthermore DMXB, a selective alpha7nAChR agonist, mimicked the PREGS effect on SIGD and antagonized the effect of PREGS. The presynaptic effect of PREGS was partially attenuated by the L-type Ca(2+) channel (VGCC) blocker nifedipine. Based on these findings, we proposed a novel mechanism underlying the facilitated synaptic transmission by PREGS: this neurosteroid sensitizes presynaptic alpha7nAChR that is followed by an activation of L-type VGCC to increase the presynaptic glutamate release.     

Studies of antidromically identified neurosecretory cells of the hypothalamus by intracellular and extracellular recordings

1. Neurosecretory neurones in supraoptic (SON) and paraventricular (PVN) nuclei of the hypothalamus of cats, anaesthetized with chloralose, and dogs, anaesthetized with Nembutal, were studied. These neurosecretory neurones were identified by action potentials evoked antidromically following stimulation of the posterior lobe of the pituitary gland. Reactions of 158 such neurones in cats and 228 in dogs were analysed.2. The latencies of antidromic potentials evoked in neurosecretory neurones by posterior lobe stimulation were between 10 and 22 msec for SON and between 14 and 28 msec for PVN cells. Approximate speed of conduction in the axons was 0.4-0.9 m/sec. The absolute refractory period for the soma-dendritic (SD) spike was 5-10 msec. These cells followed repetitive stimulation up to a rate of 100/sec.A notch was generally present on the rising phase of antidromic potentials and when the antidromically conducted signal fell in the relative refractory period of the preceding response, a complete separation between this first small A-spike and later large B-spikes, probably soma-dendritic spike, frequently occurred. Thus, two responses, a small and a large, sometimes appeared with more than 10 msec intervening. When the second antidromic response fell in the absolute refractory period of the first, the B-spike was blocked and only the A-spike appeared.3. Intracellular recordings from neurosecretory cells, mainly from SON in the dog, showed that these neurones possess resting membrane potentials of 50-80 mV, and action potentials of the same magnitude.In spontaneously firing neurosecretory cells separate A- and B-spikes also occurred and could be recorded intracellularly.4. Neurosecretory cells were excited by current injected intracellularly through a micro-electrode. The rheobase was 1-10 nA. A low intensity of stimulation only induced a small A-spike, but as the current was increased the full sized spike was evoked. Application of suprathreshold depolarizing current produced repetitive discharges. The intervals between spikes shortened with an increase in applied current intensity.5. There were a few neurones excited by stimulation of the posterior pituitary whose potentials did not meet the adopted criteria of antidromic potentials. These units were not classified as neurosecretory cells. The characteristics of cells giving the atypical ;antidromic potentials' were: the neurones discharged repetitively to antidromic stimulation, but with fluctuating and very long latencies.6. Neurosecretory cells in both SON and PVN were orthodromically excited by single pulse stimulations of the septal area, mid-brain reticular formation (RF), central gray, anterior commissure and hippocampus. The orthodromic responses generally consisted of two to three spikes with latencies of 10-30 msec. Excitation was followed by an inhibition, of ;spontaneous' discharges as well as of subsequent antidromic excitation, lasting 100-500 msec. Intracellular recordings from neurosecretory cells showed that stimulations of the septal area and RF produced action potentials or EPSPs of short duration followed by long lasting IPSPs. Hyperpolarization was always longer than the preceding EPSP, and its duration was generally 80 msec. Large IPSPs of 20 mV could be recorded occasionally.7. Antidromic excitation of neurosecretory cells by stimulation of the posterior pituitary was followed by the inhibition of ;spontaneous' discharges of the cells. This inhibition usually lasted for 100 msec. A corresponding IPSP was recorded during this inhibitory phase. These findings indicate the existence of recurrent collaterals in neurosecretory cells.8. This conclusion that recurrent collaterals exist was also supported by other evidence, namely, that certain neurones were found in the SON and PVN which responded to a single pulse antidromic stimulation of the posterior pituitary with five to seven discharges at a rate of between 500 and 800/sec. Weaker stimuli produced fewer spikes. Such cells resembled in their behaviour ;Renshaw cells' of the spinal cord. RF stimulation had an inhibitory effect on some of these neurones and an excitatory effect on others.9. Neurosecretory cells in the SON and PVN were excited by osmotic stimulation. Other neurones in close proximity were also found to be osmosensitive but they were either interneurones or neurosecretory cells whose axons ended in areas other than the posterior pituitary since they were not antidromically excitable.     

Whole-cell patch-clamp recordings from identified spinal neurons in the zebrafish embryo

We describe a preparation for obtaining patch-clamp recordings from identified embryonic spinal cord interneurons, motoneurons and sensory neurons in an in vivo zebrafish preparation. This preparation is used to study the spatial and temporal patterns of spontaneous and touch-evoked electrical activity during the initial development of circuitry in the spinal cord. The combination of these physiological techniques with the powerful genetic and molecular tools available in the zebrafish has the potential to increase our understanding of the complex interactions between genes and electrical activity during the development of the vertebrate nervous system.     

Protective effects on acute hypoxic-ischemic brain damage in mfat-1 transgenic mice by alleviating neuroinflammation

Acute hypoxic-ischemic brain damage (HIBD) mainly occurs in adults as a result of perioperative cardiac arrest and asphyxia. The benefits of n-3 polyunsaturated fatty acids (n-3 PUFAs) in maintaining brain growth and development are well documented. However, possible protective targets and underlying mechanisms of mfat-1 mice on HIBD require further investigation. The mfat-1 transgenic mice exhibited protective effects on HIBD, as indicated by reduced infarct range and improved neurobehavioral defects. RNA-seq analysis showed that multiple pathways and targets were involved in this process, with the anti-inflammatory pathway as the most significant. This study has shown for the first time that mfat-1 has protective effects on HIBD in mice. Activation of a G protein-coupled receptor 120 (GPR120)-related anti-inflammatory pathway may be associated with perioperative and postoperative complications, thus innovating clinical intervention strategy may potentially benefit patients with HIBD.