Presynaptic Density Research Articles

Cellular and molecular determinants targeting the Caenorhabditis elegans PHR protein RPM-1 to perisynaptic regions.

Caenorhabditis elegans RPM-1 is a member of a conserved protein family, the PHR proteins, that includes human Pam, mouse Phr1, zebrafish Esrom, and Drosophila Highwire. PHR proteins play important roles in the development of the nervous system. In particular, mutations in rpm-1 cause a disruption of synaptic architecture, affecting the distribution of synaptic vesicles and the number of presynaptic densities. Using antibodies against RPM-1, we determined the localization of the endogenous RPM-1 protein in wild-type and in several mutants that affect synaptic development. Our analyses show that, in mature neurons, RPM-1 resides in a distinct region that is close to, but does not overlap with, the synaptic exo- and endocytosis domains. The localization of RPM-1 occurs independently of several proteins that function in the transport or assembly of synapse components, and its abundance is partially dependent on its binding partner the F-box protein FSN-1. RPM-1 has been shown to target the MAPKKK DLK-1 for degradation. We show that activated DLK-1 may be preferentially targeted for degradation. Furthermore, using transgene analysis, we identified a critical role of the conserved PHR domain of RPM-1 in its subcellular localization.

Cognitive impairment and transmitter‐specific pre‐ and postsynaptic changes in the rat cerebral cortex during ageing

Recent studies suggest that age-related cognitive decline is correlated with an excitatory-inhibitory imbalance in synaptic discharges on pyramidal neurons. This study focuses on whether ageing and cognitive status correlates with relative numbers of excitatory and inhibitory presynaptic boutons. We investigated the density of excitatory and inhibitory presynaptic inputs across several areas of the rat cerebral cortex in young and aged male Fischer 344 rats. Aged animals were segregated into aged cognitively impaired (AI) and aged cognitively unimpaired (AU) groups using the Morris water maze. We applied immunohistochemistry to reveal the majority of excitatory and inhibitory presynaptic boutons captured with confocal microscopy and quantitative image analysis. A gradual decline in the density of excitatory and inhibitory presynaptic boutons occurred from young to AU to AI animals; however, the ratios of excitatory to inhibitory presynaptic bouton densities were not significantly altered. We further investigated the density of receptor scaffolding proteins representing key excitatory and inhibitory receptor postsynaptic sites, using antibodies against specific markers of excitatory and inhibitory postsynaptic densities, respectively. Significant changes in the ratios of excitatory to inhibitory postsynaptic densities were observed only in AI compared to young rats.

Paradoxical Upregulation of Glutamatergic Presynaptic Boutons during Mild Cognitive Impairment

Synaptic integrity is now recognized as a central component of Alzheimer's disease. Surprisingly, however, the structural status of glutamatergic synapses in Alzheimer's disease is unclear, despite the fact that glutamate is the major excitatory transmitter of the CNS and has key roles in excitotoxicity and long-term potentiation. The identification of specific markers of glutamatergic neurons now allows an assessment of the structural involvement of the glutamatergic system across progressive stages of the Alzheimer's pathology, an opportunity not afforded by previously used neurochemical approaches. Glutamatergic presynaptic bouton density and dystrophic neurite abundance were quantified in midfrontal gyrus brain tissue from subjects with no cognitive impairment, mild cognitive impairment, or mild- or severe-stage Alzheimer's disease. Our study demonstrates a striking pathology-dependent pattern of glutamatergic synaptic remodeling with disease progression. Subjects with mild cognitive impairment display a paradoxical elevation in glutamatergic presynaptic bouton density, a situation akin to that observed in the cholinergic system, which then depletes and drops with disease progression. This pattern of synaptic remodeling mirrors our previous findings in transgenic animal models and is of major relevance to current transmitter-based therapeutics.

LDL receptor deficiency results in decreased cell proliferation and presynaptic bouton density in the murine hippocampus

An aberrant cholesterol metabolism in the brain may contribute to the pathogenesis of Alzheimer's disease (AD). The LDL receptor (LDLR) regulates plasma cholesterol levels and recently we and others obtained evidence that it is also involved in regulating brain cholesterol homeostasis. Moreover, we found that LDLR-deficient mice display impaired spatial memory. Because cholesterol, in part derived from cellular uptake via LDLR, is required for peripheral cell proliferation and growth, we examined the effect of absence of the LDLR on hippocampal proliferation and the density of synaptic connections. Mice deficient for the LDLR displayed a reduced number of proliferating (BrdU-labeled) cells in the hippocampus as compared to wild type control mice. In addition, the number of synaptophysin-immunoreactive presynaptic boutons in the hippocampal CA1 and the dentate gyrus (DG) areas, but not in cortical areas, was lower in the LDLR-knockout mice than in the control mice. In vitro experiments showed that LDLR activity is increased when cell growth is enhanced by the addition of N2 supplement. This further supports a role for the LDLR in the outgrowth of neurites. These findings support the notion that, similar to its role in the periphery, the LDLR is important for the cellular uptake of cholesterol in the brain and that disturbance of this process affects neuronal plasticity.

Three-Dimensional Architecture of Presynaptic Terminal Cytomatrix

Presynaptic terminals are specialized for mediating rapid fusion of synaptic vesicles (SVs) after calcium influx. The regulated trafficking of SVs likely results from a highly organized cytomatrix. How this cytomatrix links SVs, maintains them near the active zones (AZs) of release, and organizes docked SVs at the release sites is not fully understood. To analyze the three-dimensional (3D) architecture of the presynaptic cytomatrix, electron tomography of presynaptic terminals contacting spines was performed in the stratum radiatum of the rat hippocampal CA1 area. To preserve the cytomatrix, hippocampal slices were immobilized using high-pressure freezing, followed by cryosubstitution and embedding. SVs are surrounded by a dense network of filaments. A given vesicle is connected to approximately 1.5 neighboring ones. SVs at the periphery of this network are also linked to the plasma membrane, by longer filaments. More of these filaments are found at the AZ. At the AZ, docked SVs are grouped around presynaptic densities. Filaments with adjacent SVs emerge from these densities. Immunogold localizations revealed that synapsin is located in the presynaptic bouton, whereas Bassoon and CAST (ERC2) are at focal points next to the AZ. In synapsin triple knock-out mice, the number of SVs is reduced by 63%, but the size of the boutons is reduced by only 18%, and the mean distance of SVs to the AZ is unchanged. This 3D analysis reveals the morphological constraints exerted by the presynaptic molecular scaffold. SVs are tightly interconnected in the axonal bouton, and this network is preferentially connected to the AZ.

Differential Loss of Presynaptic Dopaminergic Markers in Parkinsonian Monkeys

Assessment of dopamine nerve terminal function and integrity is a strategy employed to monitor deficits in Parkinson's disease (PD) patients and in preclinical models of PD. Dopamine replacement therapies effectively replenish the diminished supply of endogenous dopamine and provide symptomatic benefit to patients. Tyrosine hydroxylase (TH), dopamine transporter (DAT), vesicular monoamine transporter 2 (VMAT2), and amino acid decarboxylase (AADC) are widely used markers of dopaminergic neurons and terminals. The present studies were initiated to: (a) assess alterations in all four markers in the MPTP primate model of dopaminergic degeneration and (b) to determine whether L-DOPA treatment may itself modulate the expression of these markers. MPTP treatment induced a significant decline of dopaminergic immunoreactive fiber and terminal density in the basal ganglia. The amount of reduction varied between markers. The rank order of presynaptic marker loss, from most to least profound reduction, was TH > VMAT2 > DAT > AADC. Semiquantitative image analysis of relative dopaminergic presynaptic fiber and terminal density illustrated region-specific reduction of all four markers. Double immunofluorescence colocalization of two presynaptic markers on the same tissue section confirmed there was a more dramatic loss of TH than of VMAT2 or of DAT following MPTP treatment. L-DOPA treatment was associated with a significantly higher level of AADC and VMAT2 immunoreactivity in the caudate nucleus compared to placebo. These results illustrate that neurotoxic injury of the dopamine system in primates leads to altered and differential expression of presynaptic dopaminergic markers in the basal ganglia and that expression of such markers may be modulated by L-DOPA therapy. These findings have implications for the use of biomarkers of disease progression as well as for the assessment of neurorestorative strategies, such as cell replacement, for the treatment of PD.

ADAM-10 over-expression increases cortical synaptogenesis

Cortical cholinergic, glutamatergic and GABAergic terminals become upregulated during early stages of the transgenic amyloid pathology. Abundant evidence suggests that sAPPα, the product of the non-amyloidogenic α-secretase pathway, is neurotrophic both in vitro and when exogenously applied in vivo. The disintegrin metalloprotease ADAM-10 has been shown to have α-secretase activity in vivo. To determine whether sAPPα has an endogenous biological influence on cortical presynaptic boutons in vivo, we quantified cortical cholinergic, glutamatergic and GABAergic presynaptic bouton densities in either ADAM-10 moderate expressing (ADAM-10 mo) transgenic mice, which moderately overexpress ADAM-10, or age-matched non-transgenic controls. Both early and late ontogenic time points were investigated. ADAM-10 mo transgenic mice display significantly elevated cortical cholinergic, glutamatergic and GABAergic presynaptic bouton densities at the early time point (8 months). Only the cholinergic presynaptic bouton density remains significantly elevated in late-staged ADAM-10 mo transgenic animals (18 months). To confirm that the observed elevations were due to increased levels of endogenous murine sAPPα, exogenous human sAPPα was infused into the cortex of non-transgenic control animals for 1 week. Exogenous infusion of sAPPα led to significant elevations in the cholinergic, glutamatergic and GABAergic cortical presynaptic bouton populations. These results are the first to demonstrate an in vivo influence of ADAM-10 on neurotransmitter-specific cortical synaptic plasticity and further confirm the neurotrophic influence of sAPPα on cortical synaptogenesis.

P3-443: Glutamatergic presynaptic bouton density is elevated in MCI, lowered in late ad and correlates to MMSE

P3-443: Glutamatergic presynaptic bouton density is elevated in MCI, lowered in late ad and correlates to MMSE

Cytoskeleton changes following differentiation of N1E-115 neuroblastoma cell line

No systematic approach to detect expression of differentiation-related elements was published so far. The undifferentiated N1E-115 neuroblastoma cell line was switched into a neuronal phenotype by DMSO treatment and used for proteomic experiments. We used two-dimensional gel electrophoresis followed by unambiguous mass spectrometrical identification of proteins to generate a map of cytoskeleton proteins (CPs), i.e., to search for differentiation-related structures. Alpha-actin, actin-like protein 6A, gamma-tubulin complex component 2, tubulin alpha 3/alpha 7, CLIP associating protein 2, B4 integrin interactor homolog were detectable in the undifferentiated cell line exclusively and neuron-specific CPs drebrin and presynaptic density protein 95, actin-related protein 2/3, alpha and beta-centractin, PDZ-domain actin binding protein, actinin alpha 1, profilin II, ezrin, coactosin-like protein, transgelin 2, myosin light polypeptide 6, tubulin alpha 2, 6 and 7, beta tubulin (94% similar with tubulin beta-2), tubulin beta 3, tubulin tyrosine ligase-like protein 1, lamin B1 and keratin 20 were observed in the differentiated cell line only. We herein identified differentiation-related expressional patterns thus providing new evidence for the role of CPs in the process of neuronal differentiation.

Circulating insulin‐like growth factor I and functional recovery from spinal cord injury under enriched housing conditions

Voluntary locomotor training as induced by enriched housing of rats stimulates recovery of locomotion after spinal cord injury (SCI). Generally it is thought that spinal neural networks of motor- and interneurons located in the ventral and intermediate laminae within the lumbar intumescence of the spinal cord, also referred to as central pattern generators (CPGs), are the 'producers of locomotion' and play a pivotal role in the amelioration of locomotor deficits after SCI. It has been suggested that locomotor training provides locomotor-specific sensory feedback into the CPGs, which stimulates remodeling of central nervous system pathways, including motor systems. Several molecules have been proposed to potentiate this process but the underlying mechanisms are not yet known. To understand these mechanisms, we studied the role of insulin-like growth factor (IGF) I in functional recovery from SCI under normal and enriched environment (EE) housing conditions. In a first experiment, we discovered that subcutaneous administration of IGF-I resulted in better locomotor recovery following SCI. In a second experiment, detailed analysis of the observed functional recovery induced by EE revealed full recovery of hindlimb coordination and stability of gait. This EE-dependent functional recovery was attenuated by alterations in the pre-synaptic bouton density within the ventral gray matter of the lumbar intumescence or CPG area. Neutralization of circulating IGF-I significantly blocked the effectiveness of EE housing on functional recovery and diminished the EE-induced alterations in pre-synaptic bouton density within the CPG area. These results support the use of IGF-I as a possible therapeutic aid in early rehabilitation after SCI.

The amyloid pathology progresses in a neurotransmitter-specific manner

Past studies using transgenic models of early-staged amyloid pathology, have suggested that the amyloid pathology progresses in a neurotransmitter-specific manner where cholinergic terminals appear most vulnerable, followed by glutamatergic terminals and finally by somewhat more resistant GABAergic terminals. To determine whether this neurotransmitter-specific progression persists at later pathological stages, presynaptic bouton densities, and the areas of occupation and localization of plaque adjacent dystrophic neurites were quantified in 18-month-old APP K670N, M671L + PS1 M146L doubly transgenic mice. Quantification revealed that transgenic animals had significantly lower cholinergic, glutamatergic and GABAergic presynaptic bouton densities. Cholinergic and glutamatergic dystrophic neurites appear to be heavily influenced by fibrillar Aβ as both types displayed a decreasing area of occupation with respect to increasing plaque size. Furthermore, cholinergic dystrophic neurites reside in closer proximity to plaques than glutamatergic dystrophic neurites, while GABAergic dystrophic neurites were minimal regardless of plaque size. To investigate whether similarities exist in the human AD pathology, a monoclonal antibody (McKA1) against the human vesicular glutamate transporter 1 (VGluT1) was developed. Subsequent staining in AD brain tissue revealed the novel presence of glutamatergic dystrophic neurites, to our knowledge the first evidence of a structural glutamatergic deficit in the AD pathology.

Ubiquitin and Protein Turnover in Synapse Function

Enduring modification of synapses is central to long-lasting neural circuit plasticity. Such adaptations include rapid posttranslational modification of existing synaptic proteins over periods of minutes and persisting changes in the abundance of synaptic proteins over hours to days. Recently, ubiquitination and protein degradation have emerged as additional mechanisms for modifying the function and molecular composition of synapses. These recent findings raise intriguing questions as to how enduring changes at synapses are accomplished in the face of robust, ongoing molecular turnover.

GABAA Receptor Kinetics in the Cerebellar Nuclei: Evidence for Detection of Transmitter from Distant Release Sites

Neurons of the cerebellar nuclei receive GABAergic input from Purkinje cells. Purkinje boutons have several closely spaced presynaptic densities without GABA transporters, raising the possibility that neurotransmitter released by one presynaptic site diffuses to multiple postsynaptic sites. To test whether such local spillover may contribute to transmission, we studied gating of GABAA receptors at 31–33°C in cerebellar nuclear neurons acutely dissociated from mice. Currents were evoked by rapid application of long steps, brief pulses, and high-frequency trains of GABA to outside-out patches. Receptors desensitized and deactivated rapidly, and dose-response measurements estimated an EC50 of ∼30μM. From these data, a kinetic scheme was developed that replicated the recorded currents. Next, we simulated diffusion of GABA in the synaptic cleft, constrained by previous electron microscopic data, and drove the kinetic GABAA receptor model with modeled concentration transients. Simulations predicted receptor occupancies of ∼100% directly opposite the release site and ∼50% at distant postsynaptic densities, such that receptors up to 700nm from a release site opened on the timescale of the inhibitory postsynaptic currents before desensitizing. Further simulations of probabilistic release from multiple-site boutons suggested that local spillover-mediated transmission slows the onset and limits the extent of depression during high-frequency signaling.

Regulation of a DLK-1 and p38 MAP Kinase Pathway by the Ubiquitin Ligase RPM-1 Is Required for Presynaptic Development

Synapses display a stereotyped ultrastructural organization, commonly containing a single electron-dense presynaptic density surrounded by a cluster of synaptic vesicles. The mechanism controlling subsynaptic proportion is not understood. Loss of function in the C. elegans rpm-1 gene, a putative RING finger/E3 ubiquitin ligase, causes disorganized presynaptic cytoarchitecture. RPM-1 is localized to the presynaptic periactive zone. We report that RPM-1 negatively regulates a p38 MAP kinase pathway composed of the dual leucine zipper-bearing MAPKKK DLK-1, the MAPKK MKK-4, and the p38 MAP kinase PMK-3. Inactivation of this pathway suppresses rpm-1 loss of function phenotypes, whereas overexpression or constitutive activation of this pathway causes synaptic defects resembling rpm-1(lf) mutants. DLK-1, like RPM-1, is localized to the periactive zone. DLK-1 protein levels are elevated in rpm-1 mutants. The RPM-1 RING finger can stimulate ubiquitination of DLK-1. Our data reveal a presynaptic role of a previously unknown p38 MAP kinase cascade.

Microglial activation and dopamine terminal loss in early Parkinson's disease

Neuroinflammatory glial response may contribute to degenerative processes in Parkinson's disease (PD). To investigate changes in microglial activity associated with changes in the presynaptic dopamine transporter density in the PD brain in vivo, we studied 10 early-stage drug-naive PD patients twice using positron emission tomography with a radiotracer for activated microglia [(11)C](R)-PK11195 and a dopamine transporter marker [(11)C]CFT. Quantitative levels of binding potentials (BPs) of [(11)C](R)-PK11195 and [(11)C]CFT in the nigrostriatal pathway were estimated by compartment analyses. The levels of [(11)C](R)-PK11195 BP in the midbrain contralateral to the clinically affected side were significantly higher in PD than that in 10 age-matched healthy subjects. The midbrain [(11)C](R)-PK11195 BP levels significantly correlated inversely with [(11)C]CFT BP in the putamen and correlated positively with the motor severity assessed by the Unified Parkinson's Disease Rating Scale in PD. In healthy subjects, the [(11)C](R)-PK11195 BP in the thalamus and midbrain showed an age-dependent increase. In vivo demonstration of parallel changes in microglial activation and corresponding dopaminergic terminal loss in the affected nigrostriatal pathway in early PD supports that neuroinflammatory responses by intrinsic microglia contribute significantly to the progressive degeneration process of the disease and suggests the importance of early therapeutic intervention with neuroprotective drugs.

The impact of Aβ-plaques on cortical cholinergic and non-cholinergic presynaptic boutons in alzheimer's disease-like transgenic mice

A previous study in our laboratory, involving early stage, amyloid pathology in 8-month-old transgenic mice, demonstrated a selective loss of cholinergic terminals in the cerebral and hippocampal cortices of doubly transgenic (APP K670N,M671L+PSl M146L) mice, an up-regulation in the single mutant APP K670N,M671L mice and no detectable change in the PSl M146L transgenics [J Neurosci 19 (1999) 2706]. The present study investigates the impact of amyloid plaques on synaptophysin and vesicular acetylcholine transporter (VAChT) immunoreactive bouton numbers in the frontal cortex of the three transgenic mouse models previously described. When compared as a whole, the frontal cortices of transgenic and control mice show no observable differences in the densities of synaptophysin-immunoreactive boutons. An individual comparison of layer V of the frontal cortex, however, shows a significant increase in density in transgenic models. Analysis of the cholinergic system alone shows significant alterations in the VAChT-immunoreactive bouton densities as evidenced by an increased density in the single (APP K670N,M671L) transgenics and a decreased density in the doubly transgenics (APP K670N,M671L+PSl M146L). In investigating the impact of plaque proximity on bouton density at early stages of the amyloid pathology in our doubly (APP K670N,M671L+PSl M146L) transgenic mouse line, we observed that plaque proximity reduced cholinergic pre-synaptic bouton density by 40%, and yet increased synaptophysin-immunoreactive pre-synaptic bouton density by 9.5%. Distance from plaques (up to 60 μm) seemed to have no effect on bouton density; however a significant inverse relationship was visible between plaque size and cholinergic pre-synaptic bouton density. Finally, the number of cholinergic dystrophic neurites surrounding the truly amyloid, Thioflavin-S + plaque core, was disproportionately large with respect to the incidence of cholinergic boutons within the total pre-synaptic bouton population. Confocal and electron microscopic observations confirmed the preferential infiltration of dystrophic cholinergic boutons into fibrillar amyloid aggregates. We therefore hypothesize that extracellular Aβ aggregation preferentially affects cholinergic terminations prior to progression onto other neurotransmitter systems. This is supported by the observable presence of non-cholinergic sprouting, which may be representative of impending neuritic degeneration.

The degree of depression in Hamilton rating scale is correlated with the density of presynaptic serotonin transporters in 23 patients with Wilson's disease.

One of the most frequent psychiatric symptoms in patients with Wilson's disease (WD) is depression. It has been suggested that depression is associated with deficits in serotonergic neurotransmission, but, hitherto, no measurements have been performed in WD. We prospectively examined 23 adult patients (12 women, 11 men, mean age 40 years) with WD for symptoms of depression using the Hamilton rating scale for depression (HAMD). We correlated the data with the presynaptic serotonin transporter density (SERT density) in the thalamus-hypothalamus and the midbrain-pons regions measured with high resolution single-photon emission computed tomography (SPECT) 24 hours after the application of 180 MBq 2beta-carbomethoxy-3beta-(4 [(123)I]iodophenyl)tropane ( [(123)I]b-CIT). The regions of interest were determined by coregistration with a standard MRI dataset. A significant negative correlation was found between HAMD and SERT density in the thalamus-hypothalamus region (r = -0.49, p = 0.02), but not in the midbrain-pons (r = -0.31, p = 0.15). We conclude that depression in patients with Wilson's disease is correlated with alterations of serotonergic neurotransmission in the thalamus-hypothalamus region.

Convergent evidence from microdialysis and presynaptic immunolabeling for the regulation of gamma-aminobutyric acid release in the globus pallidus following acute clozapine or haloperidol administration in rats.

Antipsychotic drugs (APDs) have been primarily characterized for their effects on dopaminergic terminal regions in the brain, especially within the corpus striatum. Efferent GABA pathways are the primary outflow of striatal processing via their projections to the substantia nigra and the globus pallidus (GP). In the current study, we analyzed changes in pallidal GABA function following acute APD administration by means of in vivo microdialysis, followed by immunolabeling of presynaptic GABA terminal density in the contralateral hemisphere of the same animals. Acute administration of the atypical APD, clozapine (10 or 30 mg/kg, s.c.), produced a dose-dependent decrease in extracellular GABA. A corresponding dose-dependent increase in the density of presynaptic terminal GABA immunolabeling in the GP was found. In contrast, the typical APD, haloperidol (1 or 3 mg/kg, s.c.), had no significant effects on either measure, although a non-significant increase in extracellular GABA and decrease in the density of GABA terminal immunolabeling was noted. Paw retraction tests conducted during the time of microdialysis showed that haloperidol produced a typical pattern of highly pronounced motor impairment, while clozapine showed an atypical profile of minimal catalepsy. These complementary results obtained from in vivo neurochemistry and presynaptic neurotransmitter labeling suggest that systemic clozapine suppresses neuronal GABA release within the GP. This decrease in released pallidal GABA may play a role in the low motor side-effect liability of atypical APDs.

The organization of synaptic vesicles at tonically transmitting connections of locust visual interneurons.

Large, second-order neurons of locust ocelli, or L-neurons, make some output connections that transmit small changes in membrane potential and can sustain transmission tonically. The synaptic connections are made from the axons of L-neurons in the lateral ocellar tracts, and are characterized by bar-shaped presynaptic densities and densely packed clouds of vesicles near to the cell membrane. A cloud of vesicles can extend much of the length of this synaptic zone, and there is no border between the vesicles that are associated with neighboring presynaptic densities. In some axons, presynaptic densities are associated with discrete small clusters of vesicles. Up to 6% of the volume of a length of axon in a synaptic zone can be occupied with a vesicle cloud, packed with 4.5 to 5.5 thousand vesicles per microm(3). Presynaptic densities vary in length, from less than 70 nm to 1.5 microm, with shorter presynaptic densities being most frequent. The distribution of vesicles around short presynaptic densities was indistinguishable from that around long presynaptic densities, and vesicles were distributed in a similar way right along the length of a presynaptic density. Within the cytoplasm, vesicles are homogeneously distributed within a cloud. We found no differences in the distribution of vesicles in clouds between locusts that had been dark-adapted and locusts that had been light-adapted before fixation.

Immunological heterogeneity of synaptic and extrasynaptic forms of non-erythroid α-spectrin in the rat retina

α-, β- Spectrin is ubiquitously distributed in neuronal compartments, i.e. at the plasma membrane of neuronal cell bodies, axonal and dendritic processes, pre- and postsynaptic nerve terminals and around synaptic vesicles, ‘Brain Res. Bull. 50 (1999) 345’. Ultrastructural analyses in the rat retina using the monoclonal antibody AA6, ‘Hear. Res. 43 (1990) 199, against non-erythroid α-spectrin, αSPII (spectrin nomenclature according to ‘Blood 81 (1993) 3173’), revealed that the antibody intensively labeled the cytoplasmic face of the plasma membrane in virtually all neuronal processes. However, no significant immunolabel was observed at the presynaptic plasma membrane, around synaptic vesicles, at presynaptic densities and synaptic ribbons. Therefore, synaptic non-erythroid α-spectrin differs immunologically from extrasynaptic non-erythroid α-spectrin. This heterogeneity might contribute to the generation of distinct retinal microdomains.