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The mammalian hippocampus exhibits spontaneous sharp wave events (1-30 Hz) with an often-present superimposed fast ripple oscillation (120-220 Hz) to form a sharp wave ripple (SWR) complex. During slow-wave sleep or quiet restfulness, SWRs result from the sequential spiking of hippocampal cell assemblies initially activated during learned or imagined experiences. Additional cortical/subcortical areas exhibit SWR events that are coupled to hippocampal SWRs, and studies in mammals suggest that coupling may be critical for the consolidation and recall of specific memories. In the present study, we have examined juvenile male and female zebrafish and show that SWR events are intrinsically generated and maintained within the telencephalon and that their hippocampal homolog, the anterodorsolateral lobe (ADL), exhibits SW events with ∼9% containing an embedded ripple (SWR). Single-cell calcium imaging coupled to local field potential recordings revealed that ∼10% of active cells in the dorsal telencephalon participate in any given SW event. Furthermore, fluctuations in cholinergic tone modulate SW events consistent with mammalian studies. Moreover, the basolateral amygdala (BLA) homolog exhibits SW events with ∼5% containing an embedded ripple. Computing the SW peak coincidence difference between the ADL and BLA showed bidirectional communication. Simultaneous coupling occurred more frequently within the same hemisphere, and in coupled events across hemispheres, the ADL more commonly preceded BLA. Together, these data suggest conserved mechanisms across species by which SW and SWR events are modulated, and memories may be transferred and consolidated through regional coupling., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

Memory disruption in mild cognitive impairment (MCI) and Alzheimer's disease (AD) is poorly understood, particularly at early stages preceding neurodegeneration. In mouse models of AD, there are disruptions to sharp wave ripples (SWRs), hippocampal population events with a critical role in memory consolidation. However, the microcircuitry underlying these disruptions is under-explored. We tested whether a selective reduction in parvalbumin-expressing (PV) inhibitory interneuron activity underlies hyperactivity and SWR disruption. We employed the 5xFAD model of familial AD crossed with mouse lines labeling excitatory pyramidal cells (PCs) and inhibitory PV cells. We observed a 33% increase in frequency, 58% increase in amplitude, and 8% decrease in duration of SWRs in ex vivo slices from male and female three-month 5xFAD mice versus littermate controls. 5xFAD mice of the same age were impaired in a hippocampal-dependent memory task. Concurrent with SWR recordings, we performed calcium imaging, cell-attached, and whole-cell recordings of PC and PV cells within the CA1 region. PCs in 5xFAD mice participated in enlarged ensembles, with superficial PCs (sPCs) having a higher probability of spiking during SWRs. Both deep PCs (dPCs) and sPCs displayed an increased synaptic E/I ratio, suggesting a disinhibitory mechanism. In contrast, we observed a 46% spike rate reduction during SWRs in PV basket cells (PVBCs), while PV bistratified and axo-axonic cells were unimpaired. Excitatory synaptic drive to PVBCs was selectively reduced by 50%, resulting in decreased E/I ratio. Considering prior studies of intrinsic PV cell dysfunction in AD, these findings suggest alterations to the PC-PVBC microcircuit also contribute to impairment. SIGNIFICANCE STATEMENT We demonstrate that a specific subtype of inhibitory neuron, parvalbumin-expressing (PV) basket cells, have selectively reduced activity in a model of Alzheimer's disease (AD) during activity critical for the consolidation of memory. These results identify a potential cellular target for therapeutic intervention to restore aberrant network activity in early amyloid pathology. While PV cells have previously been identified as a potential therapeutic target, this study for the first time recognizes that other PV neuronal subtypes, including bistratified and axo-axonic cells, are spared. These experiments are the first to record synaptic and spiking activity during sharp wave ripple (SWR) events in early amyloid pathology and reveal that a selective decrease in excitatory synaptic drive to PV basket cells (PVBCs) likely underlies reduced function., (Copyright © 2020 the authors.)

Emerging evidence suggests that there is a reduction in overall cortical excitatory to inhibitory balance in major depressive disorder (MDD), which afflicts ∼14%-20% of individuals. Reduced pyramidal cell arborization occurs with stress and MDD, and may diminish excitatory neurotransmission. Enhanced deposition of perineuronal net (PNN) components also occurs with stress. Since parvalbumin-expressing interneurons are the predominant cell population that is enveloped by PNNs, which enhance their ability to release GABA, excess PNN deposition likely increases pyramidal cell inhibition. In the present study, we investigate the potential for matrix metalloprotease-9 (MMP-9), an endopeptidase secreted in response to neuronal activity, to contribute to the antidepressant efficacy of the serotonin/norepinephrine reuptake inhibitor venlafaxine in male mice. Chronic venlafaxine increases MMP-9 levels in murine cortex, and increases both pyramidal cell arborization and PSD-95 expression in the cortex of WT but not MMP-9-null mice. We have previously shown that venlafaxine reduces PNN deposition and increases the power of ex vivo γ oscillations in conventionally housed mice. γ power is increased with pyramidal cell disinhibition and with remission from MDD. Herein we observe that PNN expression is increased in a corticosterone-induced stress model of disease and reduced by venlafaxine. Compared with mice that receive concurrent venlafaxine, corticosterone-treated mice also display reduced ex vivo γ power and impaired working memory. Autopsy-derived PFC samples show elevated MMP-9 levels in antidepressant-treated MDD patients compared with controls. These preclinical and postmortem findings highlight a link between extracellular matrix regulation and MDD. SIGNIFICANCE STATEMENT Reduced excitatory neurotransmission occurs with major depressive disorder, and may be normalized by antidepressant treatment. Underlying molecular mechanisms are, however, not well understood. Herein we investigate a potential role for an extracellular protease, released from neurons and known to play a role in learning and memory, in antidepressant-associated increases in excitatory transmission. Our data suggest that this protease, matrix metalloprotease-9, increases branching of excitatory neurons and concomitantly attenuates the perineuronal net to potentially reduce inhibitory input to these neurons. Matrix metalloprotease-9 may thus enhance overall excitatory/inhibitory balance and neuronal population dynamics, which are important to mood and memory., (Copyright © 2020 the authors.)

Mutations in the gene encoding TDP-43-the major protein component of neuronal aggregates characteristic of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) with ubiquitin-positive inclusion bodies-have been linked to familial forms of both disorders. Aggregates of TDP-43 in cortical and spinal motorneurons in ALS, or in neurons of the frontal and temporal cortices in FTLD, are closely linked to neuron loss and atrophy in these areas. However, the mechanism by which TDP-43 mutations lead to neurodegeneration is unclear. To investigate the pathogenic role of TDP-43 mutations, we established a model of TDP-43 proteinopathies by expressing fluorescently tagged wild-type and mutant TDP-43 in primary rat cortical neurons. Expression of mutant TDP-43 was toxic to neurons, and mutant-specific toxicity was associated with increased cytoplasmic mislocalization of TDP-43. Inclusion bodies were not necessary for the toxicity and did not affect the risk of cell death. Cellular survival was unaffected by the total amount of exogenous TDP-43 in the nucleus, but the amount of cytoplasmic TDP-43 was a strong and independent predictor of neuronal death. These results suggest that mutant TDP-43 is mislocalized to the cytoplasm, where it exhibits a toxic gain-of-function and induces cell death.

Netrins are an important family of axon guidance cues. Here, we report that netrin-1 induces tyrosine phosphorylation of p130(CAS) (Crk-associated substrate). Our biochemical studies indicate that p130(CAS) is downstream of the Src family kinases and upstream of the small GTPase Rac1 and Cdc42. Inhibition of p130(CAS) signaling blocks both the neurite outgrowth-promoting activity and the axon attraction activity of netrin-1. p130(CAS) RNA interference inhibits the attraction of commissural axons in the spinal cord by netrin-1 and causes defects in commissural axon projection in the embryo. These results demonstrate that p130(CAS) is a key component in the netrin signal transduction pathway and plays an important role in guiding commissural axons in vivo.

Mutations in human PRPF31 gene have been identified in patients with autosomal dominant retinitis pigmentosa (adRP). To begin to understand mechanisms by which defects in this general splicing factor cause retinal degeneration, we examined the relationship between PRPF31 and pre-mRNA splicing of photoreceptor-specific genes. We used a specific anti-PRPF31 antibody to immunoprecipitate splicing complexes from retinal cells and identified the transcript of rhodopsin gene (RHO) among RNA species associated with PRPF31-containing complexes. Mutant PRPF31 proteins significantly inhibited pre-mRNA splicing of intron 3 in RHO gene. In primary retinal cell cultures, expression of the mutant PRPF31 proteins reduced rhodopsin expression and caused apoptosis of rhodopsin-positive retinal cells. This primary retinal culture assay provides an in vitro model to study photoreceptor cell death caused by PRPF31 mutations. Our results demonstrate that mutations in PRPF31 gene affect RHO pre-mRNA splicing and reveal a link between PRPF31 and RHO, two major adRP genes.

Spiral waves are a basic feature of excitable systems. Although such waves have been observed in a variety of biological systems, they have not been observed in the mammalian cortex during neuronal activity. Here, we report stable rotating spiral waves in rat neocortical slices visualized by voltage-sensitive dye imaging. Tissue from the occipital cortex (visual) was sectioned parallel to cortical lamina to preserve horizontal connections in layers III-V (500-mum-thick, approximately 4 x 6 mm(2)). In such tangential slices, excitation waves propagated in two dimensions during cholinergic oscillations. Spiral waves occurred spontaneously and alternated with plane, ring, and irregular waves. The rotation rate of the spirals was approximately 10 turns per second, and the rotation was linked to the oscillations in a one-cycle- one-rotation manner. A small (<128 mum) phase singularity occurred at the center of the spirals, about which were observed oscillations of widely distributed phases. The phase singularity drifted slowly across the tissue ( approximately 1 mm/10 turns). We introduced a computational model of a cortical layer that predicted and replicated many of the features of our experimental findings. We speculate that rotating spiral waves may provide a spatial framework to organize cortical oscillations.

Although neuronal migration is an essential process in development, how neural precursors reach their final destination in the nervous system is not well understood. Secreted molecules that are known to be involved in axon guidance are likely to play important roles in regulating neuronal migration, but an important issue that remains unclear is whether such molecules act as directional guidance cues or as motility regulators in neuronal migration. The secreted protein Slit was initially suggested to be a repellent for migrating neurons (Wu et al., 1999). However, it was concluded recently that Slit plays an inhibitory rather than a repulsive role in neuronal migration (Mason et al., 2001). We have developed a series of assays that allow us to differentiate between repulsive and inhibitory effects of secreted molecules, and we demonstrate that Slit is a repellent capable of reversing the direction of neurons migrating either in culture or in their native pathways. We also show that although Slit reduces migratory speed under certain conditions, it can function as a repellent without concurrent inhibition of neuronal migration. This is the first study to clearly demonstrate that migrating neurons can be directionally guided by secreted molecules. These findings provide a basis to understand the physiological roles of secreted molecules in the developing nervous system and have implications on how they could be applied therapeutically. Our results also indicate that it should be possible to determine the specific action of other molecules as directional guidance cues or as motility regulators of cell migration.

During development, mitral cells, the major output neurons of the olfactory bulb, project their axons caudolaterally into the telencephalon and form the lateral olfactory tract (LOT). Two types of guidance cues have been suggested for this projection. First, a long-range factor Slit, which is secreted from the septum, repels mitral cell axons into a caudolateral direction. Second, the pathway of mitral cell axons contains a subset of neurons designated as lot cells, which guide the axons through short-range interactions. It is not clear how these two guidance cues relate to each other and how they share the physiological roles. Here we examined the behavior of mitral cell axons in organotypic culture on ectopic application of Slit and inhibition of endogenous Slit signaling. The results suggested that the short-range guidance cue in the LOT pathway functions independently from Slit. Furthermore, our results showed that removal of the septum and inhibition of Slit signaling did not affect the projection of mitral cell axons. Although the septum and exogenous Slit can repel olfactory bulb axons, our results cast doubts on the physiological relevance of the septum and endogenous Slit in guiding the projection of mitral cell axons.

The Slit proteins are a new family of secreted guidance cues involved in axon guidance and neuronal migration. Each mammalian Slit protein contains >1400 amino acid residues, with four leucine-rich regions (LRRs), nine epidermal growth factor repeats, a laminin G domain, and a C-terminal cysteine-rich domain. A receptor for Slit is the transmembrane protein Roundabout (Robo), whose extracellular part contains five Ig domains and three fibronectin type III repeats. We report here that the LRRs in Slit are sufficient for binding to the Ig domains of Robo. Mutant forms of Slit containing only the LRRs function as chemorepellents for axons projecting from the olfactory bulb both in vitro and in the telencephalon. The LRRs can repel neurons migrating from the anterior subventricular zone (SVZa) to the olfactory bulb in brain slices isolated from neonatal rodents. However, the LRRs do not show repulsive effects on the SVZa neurons migrating in collagen gels. Our results indicate that the same LRRs are sufficient for guiding both axon projection and neuronal migration and suggest that the other regions in the Slit proteins may be involved in regulating the diffusion and distribution of the Slit proteins. The fact that the same domains are involved in guiding axon projection and neuronal migration further strengthens the idea of a conserved guidance mechanism for these important processes.

Population activity in the cortex is poorly understood. In this report we use voltage-sensitive dye imaging to examine the spatiotemporal patterns of a 7-10 Hz oscillation in neocortical slices from rat somatosensory areas. This oscillation appeared as a component of spontaneous epochs when the preparation was bathed in low [Mg] artificial CSF (ACSF) (Silva et al., 1991). Each epoch started with a synchronized spike, and 3-200 cycles of oscillation emerged afterward. Voltage-sensitive dye imaging revealed that the oscillations in the local field potential recordings were actually caused by a propagating population activation. This activation propagated in a relatively uniform size (not expanding). We call this confined, propagating activation a "dynamic ensemble." During each oscillation cycle, one (occasionally two) dynamic ensemble(s) appeared in the slice and was sustained for 60-200 msec. Dynamic ensembles propagated at approximately 30 mm/sec; the activity could propagate in both directions in cortical slices. The propagation consisted in part of "jumps," the locations of which were not fixed. Dynamic ensembles were distinguishable from the epileptiform spikes that occurred in low [Mg] ACSF. Population events similar to dynamic ensembles were also evoked under conditions of unaltered excitability (slice in normal ACSF) by electrical stimulation that activated a low density of neurons in a large area. Our data suggest that self-sustained, spatially confined, and propagating dynamic ensembles might be related to the epoch oscillations in somatosensory cortex seen in vivo (Nicolelis et al., 1995) and thus resemble one form of population activation in the neocortex.

Mutations in presenilin genes PS1 and PS2 account for approximately 50% of early-onset familial Alzheimer's disease (FAD). The PS1 and PS2 genes encode highly homologous transmembrane proteins related to the Caenorhabditis elegans sel-12 and spe-4 gene products. A hydrophilic loop region facing the cytoplasmic compartment is likely to be functionally important because at least 14 mutations in FAD patients have been identified in this region. We report here that the loop regions of PS1 and PS2 interact with nonmuscle filamin (actin-binding protein 280, ABP280) and a structurally related protein (filamin homolog 1, Fh1). Overexpression of PS1 appears to modify the distribution of ABP280 and Fh1 proteins in cultured cells. A monoclonal antibody recognizing ABP280 and Fh1 binds to blood vessels, astrocytes, neurofibrillary tangles, neuropil threads, and dystrophic neurites in the AD brain. Detection of ABP280/Fh1 proteins in these structures suggests that these presenilin-interacting proteins may be involved in the development of AD and that interactions between presenilins and ABP280/Fh1 may be functionally significant. The ABP280 gene is located on the human X chromosome, whereas the newly identified Fh1 gene maps to human chromosome 3. These results provide a new basis for understanding the function of presenilin proteins and further implicate cytoskeletal elements in AD pathogenesis.

Taurine is known to be involved in many important physiological functions. Here we report that both in vivo and in vitro the taurine-synthesizing enzyme in the brain, namely cysteine sulfinic acid decarboxylase (CSAD), is activated when phosphorylated and inhibited when dephosphorylated. Furthermore, protein kinase C and protein phosphatase 2C have been identified as the enzymes responsible for phosphorylation and dephosphorylation of CSAD, respectively. In addition, the effect of neuronal depolarization on CSAD activity and 32P incorporation into CSAD in neuronal cultures is also included. A model to link neuronal excitation and CSAD activation by a Ca2+-dependent protein kinase is proposed.

We examined two aspects of the response to siphon stimulation in an attempt to test the hypothesis that the Aplysia CNS functions as a distributed system. First, we estimated the number of central neurons that respond to a light touch to the siphon skin. We made voltage-sensitive dye recordings from the abdominal, pleural, pedal, and cerebral ganglia. From these recordings we estimated that 220 abdominal neurons, 110 pleural neurons, and 650 pedal neurons were affected by the light touch. Thus, the information about this mild and localized stimulus is very widely distributed within the Aplysia CNS. This result allows the possibility that the Aplysia CNS functions as a distributed system. If only a small number of neurons had responded to the touch, it would have supported the conclusion that the gill-withdrawal reflex could be generated by a small, dedicated circuit. Second, we searched for correlations between the spike times of the individual abdominal ganglion neurons. Two time scales were examined: a millisecond time scale corresponding to the duration of a fast synaptic potential and a seconds time scale corresponding to the duration of the gill-withdrawal movement. Neuron pairs with highly correlated spike activity on a millisecond time scale must be connected by (or have a common input that uses) relatively powerful, fast, excitatory synapses. We expected that this kind of synaptic interaction would be relatively rare in nervous systems that functioned in a distributed manner. Indeed, only 0.3% of the neuron pairs had correlation coefficients of 0.15 or greater. These correlations accounted for approximately 2% of the action potentials generated in response to siphon stimulation. Thus, large, fast excitatory synaptic interactions appear to be relatively unimportant. This result is consistent with the hypothesis that the abdominal ganglion functions as a distributed system. When the longer time scale was used for the cross-correlograms, a large fraction of the cell pairs had correlated activity because many neurons are activated by the stimulus. It was not possible to interpret the slow correlations in terms of actual synaptic interactions between individual neurons. Our results are consistent with the possibility that the abdominal ganglion functions in a distributed manner. However, this evaluation is indirect and thus only tentative conclusions can be drawn. Evidence from several sources suggests that the neuronal interactions for generating the Aplysia gill-withdrawal reflex are complex.

What is the internal noise in a nervous system? We studied this question by determining the trial-to-trial consistency of the neuronal response in the abdominal ganglion of Aplysia californica. Because our voltage-sensitive dye recordings detected the spike activity from a large fraction of the neurons in the ganglion, these results provide a reasonably complete characterization of the consistency of the response to a sensory stimulus. The consistency of each neuron was evaluated by the number and timing of spikes in the response. The variability in the spike count was described using the coefficient of variation. The spike count variations follow a Poisson distribution, indicating that most of these variations were the result of a random process. For each neuron the reliability of the response to touch was measured in two ways; both measures indicated a broad distribution of reliabilities within the neuron population. The time of the maximum response also varied substantially in some animals. These timing variations were in part due to random processes and in part due to systematic effects (changes in activity of many neurons that were highly correlated). The time course of the activity of individual neurons was compared with the time course of the gill withdrawal. In some animals the activity of individual neurons was only poorly correlated with the behavior; in contrast, the summed activity of groups of neurons matched the behavior quite well. This implies that the behavioral output of the system may be a distributed combination of the activity of many neurons. The differences between animals were substantially larger than the trial-to-trial differences in one animal. The responses made by different preparations differed along many dimensions.

Global observations of neuronal response in the Aplysia abdominal ganglion were made during habituation of the gill withdrawal reflex using voltage-sensitive dye recording. This technique makes it possible to measure the spike activity of 30-50% of the 1000 neurons present in the ganglion. Our experiments address the issue of how habituation is expressed in the activity of the population of neurons responding to siphon stimulation. Several classes of neurons exhibited characteristically distinct responses to the stimuli and to habituation training. One class of neurons (group I) responded to the onset and offset of the sensory stimulus although they are probably not primary sensory neurons. They habituate only partially when the behavioral reflex has already habituated completely. Two other classes (groups II and III) both have sustained responses to the touch, but habituate differently. Members of group III habituate completely while those in group II habituate only partially. Another class of neurons are inhibited by the stimulus (group IV). They become less inhibited after habituation. The response of both group I and group IV are new classes of response that have not been previously reported.

Optical recording methods using voltage-sensitive dyes were used to monitor activity in rat somatosensory cortex. We measured evoked signals in response to whisker stimulation before (control) and after the addition of the epileptogenic agent, bicuculline, and also detected spontaneous interictal events that occurred after bicuculline. Bicuculline led to an increase in the size, duration, cortical extent, and, surprisingly, the latency of the evoked responses. These enhanced evoked responses appeared to originate in the region of the control response and propagate outward. In contrast, the spontaneous signals appeared to originate at random cortical positions and had a more variable cortical extent. A transition signal measured just after the addition of bicuculline was larger than the control response but localized and rapid in time course. In most cases, there was a good correlation between the optical recordings and field potential measurements made with a ball electrode on the cortical surface, but there were occasional instances where the optical signal disappeared while the ball electrode signal was unchanged.

Immunocytochemical techniques were used to examine the synaptic relations of inhibitory interneurons in the developing rabbit hippocampus. Glutamic acid decarboxylase (GAD), the synthesizing enzyme for the inhibitory neurotransmitter GABA, was found in interneurons of immature (8 d old) as well as mature (30 d old) tissue. GAD-immunoreactivity was seen in somata, dendrites, and axon terminals of interneurons at both ages. Electron-microscopic examination revealed that GAD-positive "terminals" in immature tissue were often not associated with the usual synaptic specializations, but were rather in simple apposition to the "postsynaptic" element. In mature tissue, GAD-positive terminals made symmetric contacts primarily with pyramidal cell somata, initial segments, and proximal dendrites. In addition, GAD-positive terminals synapsed onto both GAD-positive and GAD-negative interneuron profiles.

The coexistence of gamma-aminobutyric acid (GABA), glutamate decarboxylase (GAD), and cholecystokinin (CCK)- or somatostatin-immunoreactive material in the same neurons was studied in the hippocampus and visual cortex of the cat. One-micrometer-thick serial sections of the same neuron were reacted to reveal different antigens by the unlabeled antibody enzyme method. All CCK- and somatostatin-immunoreactive neurons in the cortex and all CCK-immunoreactive and the majority of somatostatin-immunoreactive neurons in the hippocampus that could be examined in serial sections were also immunoreactive for GABA. In neurons that were immunoreactive for GAD it was often possible to demonstrate immunoreactivity for one of the peptides as well as for GABA. GABA-immunoreactive neurons, as revealed by an antiserum to GABA, were present in all layers of the cortex and hippocampus, and their shape, size, and distribution were similar to GAD-immunoreactive neurons. All GAD-immunoreactive neurons were also positive for GABA, but the latter staining revealed additional neurons. CCK/GABA- and somatostatin/GABA-immunoreactive neurons were present mainly in layers II and upper III and in layers V and VI in the visual cortex. CCK/GABA-immunoreactive neurons were most frequently present in the strata oriens, pyramidale, and moleculare of the hippocampus and in the polymorph cell layer of the dentate gyrus. Somatostatin/GABA-immunoreactive neurons were localized mainly in the stratum oriens and in the hilus of the fascia dentata. The two peptides could not be found in the same neuron. The majority of neurons that were GABA immunoreactive did not stain for either peptide. The presence of CCK- and somatostatin-immunoreactive material in GABAergic cortical neurons raises the possibility that neuroactive peptides affect GABAergic neurotransmission.

Neurons immunoreactive for the enzyme adenosine deaminase (ADA) in the posterior basal hypothalamus of the rat have a distribution pattern similar to those immunoreactive for histidine decarboxylase (HDC) and are particularly numerous in the tuberal (TM), caudal (CM) and postmammillary caudal (PCM) hypothalamic magnocellular nuclei which harbor neurons containing glutamic acid decarboxylase (GAD). The extent to which these enzymes coexist within neurons of these hypothalamic regions was examined using either serial sections or simultaneous immunostaining for ADA and HDC or GAD in the same section. Analysis of serial sections revealed neuronal coexistence of ADA with HDC or GAD in both TM and CM. In addition some neurons in CM, the only area examined for triple coexistence, were found to contain all three enzymes. In sections processed for ADA simultaneously with HDC or GAD, nearly all ADA-immunoreactive neurons in TM, CM, and PCM as well as those scattered between these nuclei were found to contain HDC, and nearly all contained GAD. Exceptions to this, however, were small cells located lateral to the posterior arcuate nucleus, which appeared to contain ADA but not HDC, and large neurons located at the anterior extreme of TM, which appeared to contain ADA but not GAD. The relatively few ADA- compared with GAD-containing neural systems in brain, together with the presence of ADA in GAD-containing hypothalamic magnocellular neurons, which appear to have widespread projections throughout the brain, indicate that ADA may be a convenient immunohistochemical marker for anatomical investigations of these projections.(ABSTRACT TRUNCATED AT 250 WORDS)

Two monoclonal antibodies against fixative-modified taurine, Tau1 and Tau2, were produced, characterized, and used in the present study to analyze the distribution of taurine in the cerebellum of the rat. In addition, immunohistochemical colocalization experiments were performed to determine whether cerebellar neurons contain both taurine and its synthesizing enzyme, cysteine sulfinic acid decarboxylase (CSADC). In ELISAs, both Tau1 and Tau2 displayed high affinities for taurine conjugated to various carrier proteins and possessed some cross-reactivity for other amino acids which are present in lower concentrations in the brain than taurine. Tau2 was found to recognize only taurine and hypotaurine when paraformaldehyde was used to fix the amino acids to carrier proteins. With the use of glutaraldehyde fixation, Tau1 cross-reacted with conjugates of beta-alanine and hypotaurine and Tau2 cross-reacted strongly with conjugates of cysteic acid and hypotaurine and weakly with cysteine sulfinic acid. Despite different cross-reactivities, Tau1 and Tau2 exhibited almost identical patterns of neuronal staining in bands of Purkinje cells in the cerebellum. Staining of Purkinje cell dendrites was more prominent than staining of the soma. Light immunoreactivity was present in Golgi, stellate, and basket cells. A scattered population of granule cells displayed taurine-like immunoreactivity at the electron microscopic level. Immunostaining was identified in some terminals in the Purkinje cell layer and in a limited number of mossy fibers. Tau2-like immunoreactivity was colocalized with CSADC-like immunoreactivity in the cerebellar neurons described above. These immunoreactive cells may represent a subpopulation of neurons that contain a higher concentration of taurine than neighboring cells due to their ability to synthesize taurine. The intense immunoreactive staining of Purkinje cell dendrites provides support for the hypothesis that calcium-dependent release of taurine in the cerebellum may originate primarily from dendritic rather than synaptic processes and suggests a neuromodulator role for taurine in the cerebellum.

An immunocytochemical study using anti-GAD serum was performed to examine the plastic changes of GABAergic inhibitory synapses in the red nucleus (RN) after lesions of the nucleus interpositus (IP) of the cat. Light-microscopic analyses revealed that 20-175 d after the unilateral lesion of the IP, somatic profiles of large neurons in the magnocellular RN contralateral to the lesion were more densely covered with GAD-immunoreactive puncta than those in the ipsilateral RN. Electron-microscopic analyses demonstrated that the GAD-immunoreactive puncta observed with the light microscope were synaptic terminals and that the number of GAD-immunoreactive synaptic terminals per unit length of somatic membrane of RN neurons was increased on the deafferented side. The GAD-immunoreactive terminals on somata of RN neurons made symmetric synaptic contacts with somatic membranes on both the deafferented and control sides. The number of immunoreactive synapses on somata of RN neurons was markedly increased on the deafferented side following IP lesion, whereas that of the unlabeled asymmetric synapses was decreased. These observations indicate that new GABAergic synapses were formed on somata of RN neurons after deafferentation from the IP.

In order to examine whether L-aspartate and L-glutamate are photoreceptor transmitters in monkey retina, we have carried out two different studies: an autoradiographic localization of the high-affinity uptake sites for aspartate (Asp) and glutamate (Glu), and an immunocytochemical localization of the biosynthetic enzyme, L-aspartate aminotransferase (AAT). Our results show that L-Glu is taken up by a sodium-dependent, high-affinity uptake system with a Km = 8 +/- 3.5 X 10(-6) M and a Vmax = 48 +/- 14 X 10(-12) mol/min/mg protein; 3H-L-Glu or 3H-L- and D-Asp are taken up and accumulated by rod somata and inner segments in retinas incubated at 37 degrees C or at ambient temperature; cones accumulate 3H-L-Glu at ambient temperature but not at 37 degrees C. Neither 3H-L-Asp nor D-Asp is accumulated by cones at either temperature; rods and cones show the same labeling pattern irrespective of whether they are located in the central or peripheral retina; and antiserum to AAT preferentially stains the cone somata, inner segments, and synaptic pedicles, while the outer segments are negative. Both foveal and peripheral cones are strongly reactive, but rods show little reactivity. These findings point to the existence of major differences between rods and cones in the uptake and metabolism of L-Asp and L-Glu and are consistent with L-Glu being a cone transmitter in the monkey retina.

A combination of optical and electrode recording methods was used to obtain an overview of the neuron activity in the Aplysia abdominal ganglion in response to a light touch to the siphon skin. Spike activity was detected in up to 150 different neurons. Habituation and sensitization of the gill-withdrawal reflex was accompanied by large changes in the number of activated neurons. It is likely that these recordings are incomplete; the actual number of activated neurons is estimated to be about 300 in the acutely sensitized preparation. While we presume that not all 300 of these neurons are involved in the gill-withdrawal reflex, the number of neurons is so large that it may be difficult to determine the role of each activated neuron with presently available experimental tools.