Small Cytoplasmic Granules Research Articles

Single mRNP Analysis Reveals that Small Cytoplasmic mRNP Granules Represent mRNA Singletons

Small cytoplasmic mRNP granules are implicated in mRNA transport, translational control, and decay. Using super-resolution microscopy and fluorescence correlation spectroscopy, we analyzed the molecular composition and dynamics of single cytoplasmic YBX1_IMP1 mRNP granules in live cells. Granules appeared elongated and branched, with patches of IMP1 and YBX1 distributed along mRNA, reflecting the attachment of the two RNA-binding proteins in cis. Particles form at the nuclear pore and do not associate with translating ribosomes, so the mRNP is a repository for mRNAs awaiting translation. In agreement with the average number of mRNA-binding sites derived from crosslinked immunoprecipitation (CLIP) analyses, individual mRNPs contain 5-15 molecules of YBX1 and IMP1 and a single poly(A) tail identified by PABPC1. Taken together, we conclude that small cytoplasmic mRNP granules are mRNA singletons, thus depicting the cellular transcriptome. Consequently, expression of functionally related mRNAs in RNA regulons is unlikely to result from coordinated assembly.

Deubiquitylase USP9X maintains centriolar satellite integrity by stabilizing pericentriolar material 1 protein.

Centriolar satellites are small cytoplasmic granules that play important roles in regulating the formation of centrosomes and primary cilia. Ubiquitylation of satellite proteins, including the core satellite scaffold protein pericentriolar material 1 (PCM1), regulates centriolar satellite integrity. Currently, deubiquitylases that control centriolar satellite integrity have not been identified. In this study, we find that the deubiquitylase USP9X binds PCM1, and antagonizes PCM1 ubiquitylation to protect it from proteasomal degradation. Knockdown of USP9X in human cell lines reduces PCM1 protein levels, disrupts centriolar satellite particles and causes localization of satellite proteins, such as CEP290, to centrosomes. Interestingly, knockdown of mindbomb 1 (MIB1), a ubiquitin ligase that promotes PCM1 ubiquitylation and degradation, in USP9X-depleted cells largely restores PCM1 protein levels and corrects defects caused by the loss of USP9X. Overall, our study reveals that USP9X is a constituent of centriolar satellites and functions to maintain centriolar satellite integrity by stabilizing PCM1.

Localization of Barley yellow dwarf virus Movement Protein Modulating Programmed Cell Death in Nicotiana benthamiana.

Barley yellow dwarf virus (BYDV) belongs to Luteovirus and is limited only at phloem related tissues. An open reading frame (ORF) 4 of BYDV codes for the movement protein (MP) of BYDV gating plasmodesmata (PD) to facilitate virus movement. Like other Luteoviruses, ORF 4 of BYDV is embedded in the ORF3 but expressed from the different reading frame in leaky scanning manner. Although MP is a very important protein for systemic infection of BYDV, there was a little information. In this study, MP was characterized in terms of subcellular localization and programmed cell death (PCD). Gene of MP or its mutant (ΔMP) was expressed by Agroinfiltration method. MP was clearly localized at the nucleus and the PD, but ΔMP which was deleted distal N-terminus of MP showed no localization to PD exhibited the different target with original MP. In addition to PD localization, MP appeared associated with small granules in cytoplasm whereas ΔMP did not. MP associated with PD and small granules induced PCD, but ΔMP showed no association with PD and small granules did not exhibit PCD. Based on this study, the distal N-terminal region within MP is seemingly responsible for the localization of PD and the induction small granules and PCD induction. These results suggest that subcellular localization of BYDV MP may modulate the PCD in Nicotiana benthamiana.

Phagocytic activities of hemocytes from the deep-sea symbiotic mussels Bathymodiolus japonicus, B. platifrons, and B. septemdierum

Deep-sea mytilid mussels harbor symbiotic bacteria in their gill epithelial cells that are horizontally or environmentally transmitted to the next generation of hosts. To understand the immune defense system in deep-sea symbiotic mussels, we examined the hemocyte populations of the symbiotic Bathymodiolus mussel species Bathymodiolus japonicus, Bathymodiolus platifrons, and Bathymodiolus septemdierum, and characterized three types of hemocytes: agranulocytes (AGs), basophilic granulocytes (BGs), and eosinophilic granulocytes (EGs). Of these, the EG cells were the largest (diameter, 8.4–10.0 μm) and had eosinophilic cytoplasm with numerous eosinophilic granules (diameter, 0.8–1.2 μm). Meanwhile, the BGs were of medium size (diameter, 6.7–8.0 μm) and contained small basophilic granules (diameter, 0.3–0.4 μm) in basophilic cytoplasm, and the AGs, the smallest of the hemocytes (diameter, 4.8–6.0 μm), had basophilic cytoplasm lacking granules. A lectin binding assay revealed that concanavalin A bound to all three hemocyte types, while wheat germ agglutinin bound exclusively to EGs and BGs. The total hemocyte population densities within the hemolymph of all three Bathymodiolus mussel species were similar (8.4–13.3 × 105 cells/mL), and the percentages of circulating AGs, BGs, and EGs in the hemolymph of these organisms were 44.7–48.5%, 14.3–17.6%, and 34.3–41.0%, respectively. To analyze the functional differences between these hemocytes, the phagocytic activity and post-phagocytic phagosome–lysosome fusion events were analyzed in each cell type using a fluorescent Alexa Fluor® 488-conjugated Escherichia coli bioparticle and a LysoTracker® lysosomal marker, respectively. While the AGs exhibited no phagocytic activity, both types of granulocytes were phagocytic. Of the three hemocyte types, the EGs exhibited the highest level of phagocytic activity as well as rapid phagosome–lysosome fusion, which occurred within 2 h of incubation. Meanwhile, the BGs showed lower phagocytic activity and lower rates of phagosome–lysosome fusion than the EGs. These findings indicate that the two types of granulocyte play distinct roles in the defense system.

Mini-review: Eosinophils, a Useful Diagnostic Clue in Surgical Neuropathology

<p>Eosinophils are one of the polymorphonuclear granulocytes derived from bone marrow stem cells, and they contain many small cytoplasmic granules that stain bright red with eosin or brick-red with Romanowsky staining. Eosinophilic infiltration is also present in various human central nervous system (CNS) diseases such as parasitic infection, transverse myelitis, vasculitis, Langerhans cell histiocytosis, glioblastoma and etc... Due to the morphologic and functional characteristics, the presence of eosinophils in certain lesions may provide useful diagnostic clues in the right clinical setting. Consideration of this finding may facilitate the diagnosis of CNS pathologic lesions, especially in a small specimen such as a stereotactic biopsy.</p>

Intracellular distribution of amyloid beta peptide and its relationship to the lysosomal system

BackgroundAmyloid beta peptide (Aβ) is the main component of extraneuronal senile plaques typical of Alzheimer’s disease (AD) brains. Although Aβ is produced by normal neurons, it is shown to accumulate in large amounts within neuronal lysosomes in AD. We have recently shown that under normal conditions the majority of Aβ is localized extralysosomally, while oxidative stress significantly increases intralysosomal Aβ content through activation of macroautophagy. It is also suggested that impaired Aβ secretion and resulting intraneuronal increase of Aβ can contribute to AD pathology. However, it is not clear how Aβ is distributed inside normal neurons, and how this distribution is effected when Aβ secretion is inhibited.MethodsUsing retinoic acid differentiated neuroblastoma cells and neonatal rat cortical neurons, we studied intracellular distribution of Aβ by double immunofluorescence microscopy for Aβ40 or Aβ42 and different organelle markers. In addition, we analysed the effect of tetanus toxin-induced exocytosis inhibition on the intracellular distribution of Aβ.ResultsUnder normal conditions, Aβ was found in the small cytoplasmic granules in both neurites and perikarya. Only minor portion of Aβ was colocalized with trans-Golgi network, Golgi-derived vesicles, early and late endosomes, lysosomes, and synaptic vesicles, while the majority of Aβ granules were not colocalized with any of these structures. Furthermore, treatment of cells with tetanus toxin significantly increased the amount of intracellular Aβ in both perikarya and neurites. Finally, we found that tetanus toxin increased the levels of intralysosomal Aβ although the majority of Aβ still remained extralysosomally.ConclusionOur results indicate that most Aβ is not localized to Golgi-related structures, endosomes, lysosomes secretory vesicles or other organelles, while the suppression of Aβ secretion increases intracellular intra- and extralysosomal Aβ.

Concholepas hemocyanin biosynthesis takes place in the hepatopancreas, with hemocytes being involved in its metabolism

Hemocyanins are copper-containing glycoproteins in some molluscs and arthropods, and their best-known function is O(2) transport. We studied the site of their biosynthesis in the gastropod Concholepas concholepas by using immunological and molecular genetic approaches. We performed immunohistochemical staining of various organs, including the mantle, branchia, and hepatopancreas, and detected C. concholepas hemocyanin (CCH) molecules in circulating and tissue-associated hemocytes by electron microscopy. To characterize the hemocytes, we purified them from hemolymph. We identified three types of granular cells. The most abundant type was a phagocyte-like cell with small cytoplasmic granules. The second type contained large electron-dense granules. The third type had vacuoles containing hemocyanin molecules suggesting that synthesis or catabolism occurred inside these cells. Our failure to detect cch-mRNA in hemocytes by reverse transcription with the polymerase chain reaction (RT-PCR) led us to propose that hemocytes instead played a role in CCH metabolism. This hypothesis was supported by colloidal gold staining showing hemocyanin molecules in electron-dense granules inside hemocytes. RT-PCR analysis, complemented by in situ hybridization analyses with single-stranded antisense RNAs as specific probes, demonstrated the presence of cch-mRNA in the hepatopancreas; this was consistent with the specific hybridization signal and confirmed the hepatopancreas as the site of CCH synthesis. Finally, we investigated the possibility that CCH catabolism in hemocytes was involved in the host immune response and in the generation of secondary metabolites such as antimicrobial peptides and phenoloxidase.

Vascular Endothelial Growth Factor–C (VEGF-C) Expression in Normal Human Tissues

To characterize vascular endothelial growth factor-C (VEGF-C) protein expression in normal human tissues by immunohistochemistry (IHC). VEGF-C is a growth factor for lymphatic endothelial cells. VEGF-C mRNA and protein are expressed in a variety of cancerous tissues, but the localization of VEGF-C protein in many normal human tissues has not been clearly demonstrated to date. We therefore performed an immunohistochemical survey of the distribution of intracellular VEGF-C protein in a range of normal human tissue types. Five microm sections were cut from archived human tissues. Sections were dewaxed, rehydrated, and subjected to microwave pretreatment. They were incubated with VEGF-C antibody before detection with biotinylated secondary antibody using 'Elite' avidin-biotin enzyme complex and diaminobenzidine substrate. The primary antibody recognized the C-terminus of the VEGF-C propeptide that is cleaved before secretion and hence only cellular protein was detected. Negative controls used the same concentration of normal goat IgG. Staining manifested as small punctate cytoplasmic granules. Strong expression was observed in large intestine epithelium, and mammary duct epithelium, skeletal and cardiac muscle, thyroid, ovary, and the prostate. Weaker expression was also detected in the hepatocytes close to the terminal hepatic venules of the liver, vascular smooth muscle, and placenta. No expression was consistently detected in spleen or thymus. Intracellular VEGF-C protein is widely expressed in many normal human adult tissues. Its expression in cancer is not therefore per se indicative of a prolymphangiogenic change. To demonstrate the latter, a quantitative change in expression level is required.

Methylation regulates the intracellular protein-protein and protein-RNA interactions of FMRP

FMRP, the fragile X mental retardation protein, is an RNA-binding protein that interacts with approximately 4% of fetal brain mRNA. We have recently shown that a methyltransferase (MT) co-translationally methylates FMRP in vitro and that methylation modulates the ability of FMRP to bind mRNA. Here, we recapitulate these in vitro data in vivo, demonstrating that methylation of FMRP affects its ability to bind to FXR1P and regulate the translation of FMRP target mRNAs. Additionally, using double-label fluorescence confocal microscopy, we identified a subpopulation of FMRP-containing small cytoplasmic granules that are distinguishable from larger stress granules. Using the oxidative-stress induced accumulation of abortive pre-initiation complexes as a measure of the association of FMRP with translational components, we have demonstrated that FMRP associates with ribosomes during initiation and, more importantly, that methylation regulates this process by influencing the ratio of FMRP-homodimer-containing mRNPs to FMRP-FXR1P-heterodimer-containing mRNPs. These data suggest a vital role for methylation in normal FMRP functioning.

Immunohistochemical staining for adipophilin, perilipin and TIP47

Background: The presence of lipid in the cell cytoplasm is useful for supporting the diagnosis of sebaceous gland carcinoma (SGC). Currently this requires histochemical stains that are carried out on...

The U L 41 protein of herpes simplex virus 1 degrades RNA by endonucleolytic cleavage in absence of other cellular or viral proteins

The herpes simplex virus 1 ORF U(L)41 encodes a protein (virion host shutoff or vhs) associated with selective degradation of mRNA early in infection. Some mRNAs, exemplified by GAPDH or beta-actin mRNAs, are degraded rapidly. Others, for example IEX-1 mRNA, are degraded in two stages: whereas the 3' domain disappears rapidly, a large 5' domain fragment of the mRNA lingers for several hours. Still a third, exemplified by tristetraprolin mRNA, is not degraded, allowing its protein product to accumulate in infected cells. Here we report the following: (i) a GST-vhs protein produced in Escherichia coli, solubilized and purified to homogeneity acts as bona fide endoribonuclease when tested on in vitro transcribed IEX-1 probes. A GST-vhs protein in which three key vhs amino acids were replaced with alanines, solubilized and purified by the same protocol, had no enzymatic activity. (ii) The number of fragments generated by cleavage of a truncated IEX-1 RNA by vhs appears to be small; the cleavage sites are centered at or near the AU-rich elements located at the 3' untranslated region of the mRNA. A truncated RNA containing only the IEX-1 coding domain was cleaved numerous times. (iii) In cells infected at high multiplicity and exposed to a large number of particles per cell, the vhs protein accumulated within 3 h after infection, in small uniform cytoplasmic granules raising the possibility that vhs colocalizes with tristerapolin, a protein induced after infection, in structures involved in degradation of RNA.

Autophagy of amyloid beta-protein in differentiated neuroblastoma cells exposed to oxidative stress

Oxidative stress is considered important for the pathogenesis of Alzheimer disease (AD), which is characterized by the formation of senile plaques rich in amyloid beta-protein (Aβ). Aβ cytotoxicity has been found dependent on lysosomes, which are abundant in AD neurons and are shown to partially co-localize with Aβ. To determine whether oxidative stress has any influence on the relationship between lysosomes and Aβ 1–42 (the most toxic form of Aβ), we studied the effect of hyperoxia (40% versus 8% ambient oxygen) on the intracellular localization of Aβ 1–42 (assessed by immunocytochemistry) in retinoic acid differentiated SH-SY5Y neuroblastoma cells maintained in serum-free OptiMEM medium. In control cells, Aβ 1–42 was mainly localized to small non-lysosomal cytoplasmic granules. Only occasionally Aβ 1–42 was found in large (over 1 μm) lysosomal-associated membrane protein 2 positive vacuoles, devoid of the early endosomal marker rab5. These large Aβ 1–42-containing lysosomes were not detectable in the presence of serum (known to suppress autophagy), while their number increased dramatically (up to 24-fold) after exposure of cells to hyperoxia during 5 days. Activation of autophagy by hyperoxia was confirmed by transmission electron microscopy. Furthermore, an inhibitor of autophagic sequestration 3-methyladenine prevented the accumulation of Aβ 1–42-positive lysosomes due to hyperoxia. In parallel experiments, intralysosomal accumulation of Aβ 1–40 following oxidative stress has been found as well. The results suggest that Aβ can be autophagocytosed and its accumulation within neuronal lysosomes is enhanced by oxidative stress.

Evidence for perforin-like activity associated with earthworm leukocytes

Earthworm (Eisenia fetida) coelomic fluid contains several leukocytes (coelomocytes): basophils, acidophils and neutrophils as well as chloragocytes. Small coelomocytes and coelomocyte lysate are cytotoxic for the tumor cell target K562. The expression of a lytic factor was investigated by immunocytochemistry using light and transmission electron microscopy. A rat-anti-mouse-perforin-mAb labeled mainly small coelomocytes (nearly 20%) as visualized by light microscopy. TEM analysis using immunogold showed a homogenous labeling in the cytoplasm of small coelomocytes. The highest number of immunogold particles was estimated in coelomocytes with many small cytoplasmic granules. Coelomocytes with large lysosomal granules were also labeled but less intensely. No antibody binding was observed for chloragocytes either in light or electron microscopy. This suggests that the perforin-like activity is associated with only one cell type and that chloragocytes are responsible for other lytic activities. MALDI-MS revealed calreticulin usually associated with perforin in mammalian cells that mediate lysis (e.g. NK, CTL). Together, results strongly suggest the presence of putative perforin in earthworms. This in turn supports the hypothesis that perforin is a conserved component important in immune defense during evolution.

Specific staining of nonpyramidal cell populations of the cerebral cortex by lectin cytochemistry on semithin sections

The pattern of lectin labeling in the cerebral cortex of the cat was studied using semithin sections. The labeling produced by some lectins (Concanavalin A , Lens culinaris, Phaseolus vulgaris-L, Phaseolus vulgaris-E, Pisum sativum, wheat germ agglutinin, and succynilated-wheat germ) appeared inside every neuron as small cytoplasmic granules, probably corresponding to cisterns of endoplasmic reticulum and/or the Golgi complex. Lectins with affinity for α-mannosyl residues ( Pisum sativum, Lens culinaris, and Concanavalin A) stained the cell surface of a subset of cortical neurons. The labeled cells were round or polygonal, medium to large neurons present in layers II–VI, exhibiting the morphological features of nonpyramidal cells. Previous lectin studies of perineuronal nets have shown that these extracellular specializations contain N-acetylgalactosamine and N-acetylglucosamine. Our results show that mannose is also a component of perineuronal nets and that lectins specific for α-mannose can be used as tools for the cytochemical detection of a separate class of cortical neurons, which have not yet been fully characterized. In addition, some lectins ( Bandeiraea simplicifolia, Concanavalin A, Lens culinaris, Phaseolus vulgaris-L, Phaseolus vulgaris-E, Pisum sativum, and succynilated-wheat germ agglutinin) specifically labeled a population of a type of microglia-related cells known as perivascular cells. The data presented here report for the first time the selective staining of perivascular cells and further support the hypothesis that they are different from typical microglial cells.

Arylsulfatase B is present in crystalloid-containing granules of human eosinophil granulocytes.

Eosinophil granulocytes are characterized by large crystalloid-containing granules whose major contents of highly cationic proteins may play a role in allergic reactions and parasitic infections. Human eosinophils are also rich in arylsulfatase B whose enzymatic activity is localized to a population of small type cytoplasmic granules and used as a marker for such organelles. We utilized immunoelectron microscopy to investigate its subcellular distribution in human eosinophils. The arylsulfatase B antigen was found to be concentrated to both the crystalloid core and the matrix of crystalloid-containing granules as well as in small type granules. Therefore arylsulfatase seems to be present primarily in crystalloid-containing granules in a possibly inactive form (but detected by antibodies) that is converted to an enzymatically active form, e.g. during secretion and formation of small type granules which may derive from the former granules.

Ultrastructure of the peritoneal exudate cells of seawater teleosts, seabream (Sparus aurata) and sea bass (Dicentrarchus labrax)

The peritoneal exudates of seabream and sea bass consist of granulocytes, lymphocytes and macrophages. These cells show conspicuous ultrastructural differences from the same cell-types of blood and head-kidney, which have not been reported previously. Peritoneal exudate granulocytes differ from their corresponding circulating or head-kidney forms in the following way: (a) they are larger in size, and (b) their abundant cytoplasmic granules have some new ultrastructural features, and a new granule population might also be present. Likewise, lymphocytes also show a noticeable difference; they contain a sparse population of small dense cytoplasmic granules. Monocytes, macrophages, and transitional forms between these two cell-types, are also found. The percentage of peritoneal exudate cell-types is different in seabream and sea bass. Macrophages in sea bass represent the most abundant peritoneal exudate cell-type. However, seabream shows lower percentages of macrophages than granulocytes.

Development, differentiation, and proliferation of macrophages in the rat yolk sac

Immunohistochemical and immunoelectron microscopic investigation using anti-rat macrophage monoclonal antibody (mAb) RM-1 demonstrated the first emergence of immature macrophages within blood islands of fetal rat yolk sacs at fetal day 9. At fetal day 10, they matured into fetal macrophages, showed intense immunoreactivity to RM-1, developed lysosomal granules, endocytic vesicles or vacuoles, and extended fine cytoplasmic processes By the rosetting assay with IgG-coated sheep erythrocyte antibody (IgG-EA), both the immature and mature fetal macrophages showed rosette formation and phagocytosis of IgG-EA, but they were negative for peroxidase (PO) reaction. At fetal day 11, fetal macrophages were observed in the mesenchymal layer of the yolk sacs. In the yolk sacs, no promonocytes or monocytes were observed, although there was a very minor population (less than 1%) of immature myeloid cells containing a few small PO-positive cytoplasmic granules from fetal day 11 on. After combination of the vitelline vessels to the embryonic cardiovascular system, fetal macrophages appeared in embryonic rat tissues. By [ 3H]thymidine autoradiography, yolk sac macrophages were demonstrated to possess a marked proliferativc potential. These results suggest that fetal macrophages in the yolk sac differentiate from haematopoietic stem cells without passing through the promonocyte or monocyte stage, proliferate actively, and colonize the embryonic rat tissues.

Characterization of A Phenotypically and Functionally Distinct Subset of Large Granular Lymphocytes (Pit Cells) in Rat Liver Sinusoids

Pit cells, or large granular lymphocytes, with natural tumoricidal activity are found in the sinusoids of normal rat liver. Hepatic large granular lymphocytes are heterogeneous and can be subdivided into two subsets. These subsets were compared with peripheral-blood large granular lymphocytes and were found to differ phenotypically and functionally. Phenotypical differences included lower expression of the asialo-GM1 marker of natural killer cells, lower cellular density and many more small cytoplasmic granules in hepatic large, granular lymphocytes. Low-density hepatic large, granular lymphocytes were five to eight times more cytotoxic than blood large, granular lymphocytes on a per-cell basis as measured against YAC-1 cells and colon carcinoma cells. In addition, hepatic large, granular lymphocytes were able to lyse P-815 target cells, which are resistant to blood natural killer cells. Large, granular lymphocytes isolated from the liver also contained a subset with intermediate phenotypical and functional characteristics, possibly representing a transitional form between blood and "liver-specific" large, granular lymphocytes. The liver thus contains a specific population of highly activated or further differentiated large granular lymphocytes.

Maturation of the endocrine pancreas in the sea bass, Dicentrarchus labrax L. (Teleostei): An immunocytochemical and ultrastructural study : I. Glucagon-producing cells

The structure of the endocrine pancreas in the sea bass ( Dicentrarchus labrax L.) was studied with special reference to glucagon-immunoreactive cells. As described in most of the teleosts, the sea bass was found to have a diffuse pancreas. In the adult, endocrine cells were clustered in a principal islet and numerous accessory islets where the glucagon A cells were localized peripherally. Under electron microscopy, the A cells displayed a clear hyaloplasm with granules having typical spherical or polyhedral cores, as in other vertebrates. The maturation of the endocrine pancreas was monitored under rearing conditions. The endocrine pancreas appeared during the prelarval stage, 3 days after hatching, and consisted of a single cluster of morphologically similar cells, containing very small cytoplasmic granules. During the larval stage, cytodifferentiation resulted in modifications of cell shape and increased granule size. Typical granules appeared in 8-mm-long larvae. Cells immunoreactive with mammalian glucagon antibodies appeared only at the beginning of the juvenile stage (3 months/20 mm). Electron microscope observations revealed that the storage of hormone in numerous cytoplasmic granules began at this stage.

Immunohistochemical localization of GABAAreceptors in the retina of the new world primateSaimiri sciureus

A large population of amacrine cells in the retina are thought to use GABA as an inhibitory neurotransmitter in their synaptic interactions within the inner plexiform layer. However, little is known about their synaptic targets; the neurons that express the receptors for GABA have not been clearly identified. Recently, the GABAA receptor has been isolated and antibodies have been raised against it. These antibodies have proven useful for the immunocytochemical localization of the receptor, and two brief reports describing the distribution of GABAA receptor immunoreactivity in the retina have appeared (Richards et al., 1987; Mariani et al., 1987). We used a monoclonal antibody (62-3G1) against the GABAA receptor to study the retina of the New World primate Saimiri sciureus. Labeled somata were found in the inner nuclear layer (INL) and ganglion cell layer (GCL). The staining was confined to what appeared to be the cell's plasmalemma and small cytoplasmic granules. Most of the labeled neurons in the INL had small somata (5-7 microns in diameter) located at the vitreal edge of the layer. They arborized in two laminae (approximately 2 and 4) of inner plexiform layer (IPL). Ventral to the optic disc (2.5 mm) they comprised 29% of the cells present. A few of the labeled neurons appeared to be interplexiform cells or flat bipolar cells, with labeled processes that extended into both the IPL and the inner half of the outer plexiform layer. In the GCL, the labeled somata were among the largest present (13-20 microns in diameter), and 2.5 mm ventral to the optic disc they made up 15% of the cells present. Experiments in which immunoreactive somata were retrogradely labeled following the injection of fluorescent tracers into the optic tract provided a conclusive demonstration that some of the immunoreactive somata were ganglion cells. The antibody often labeled their axons in the optic fiber layer. This suggests that the GABAA receptors are transported anterogradely to the retinal terminal fields. The dendrites of the immunoreactive ganglion cells extended into the 2 laminae of labeled processes in the IPL, and their primary dendritic arbors were, at any given eccentricity, quite similar in appearance. This homogeneity suggests that they comprise a particular subset of the ganglion cells. Sections simultaneously labeled with the monoclonal antibody against the GABAA receptor and antisera against either L-glutamic acid decarboxylase (GAD) or GABA revealed that the GAD/GABA was distributed much more widely in the IPL than the GABAA receptor.(ABSTRACT TRUNCATED AT 400 WORDS)