Rebecca C. Stacy

Developmental relationship between cholinergic amacrine cell processes and ganglion cell dendrites of the mouse retina

Ganglion cells of the mammalian retina undergo structural remodeling before their dendrites are confined to functionally distinct laminas within the inner plexiform layer. It has been proposed that cholinergic amacrine cells provide laminar cues that remodel ganglion cell dendrites, because their processes stratify before those of the ganglion cells. To address this possibility, it is necessary to know whether cholinergic cells contact all or only some classes of ganglion cells during development. We, therefore, used two‐photon microscopy to simultaneously reconstruct the dendritic arbors of different classes of ganglion cells and terminal processes of cholinergic cells in neonatal mouse retina. We determined that, after birth, cholinergic cells contacted only a subset of ganglion cells. Large bistratified cells (LBCs), resembling direction selective ganglion cells in other species, had dendrites that fasciculated with the cholinergic plexuses. The LBCs received numerous presynaptic cholinergic contacts shortly after birth. In contrast, large monostratified cells (LMCs), ramifying outside the cholinergic plexuses at maturity, received few, if any, cholinergic contacts even at early stages when their dendrites overlapped with the cholinergic processes. These observations suggest that cholinergic cells provide laminar cues for only subsets of ganglion cells. They also indicate that the synaptic organization between amacrine and ganglion cells may be specified early in development. J. Comp. Neurol. 456:154–166, 2003.

Disruption and Recovery of Patterned Retinal Activity in the Absence of Acetylcholine

Many developing neural circuits generate synchronized bursting activity among neighboring neurons, a pattern thought to be important for sculpting precise neural connectivity. Network output remains relatively constant as the cellular and synaptic components of these immature circuits change during development, suggesting the presence of homeostatic mechanisms. In the retina, spontaneous waves of activity are present even before chemical synapse formation, needing gap junctions to propagate. However, as synaptogenesis proceeds, retinal waves become dependent on cholinergic neurotransmission, no longer requiring gap junctions. Later still in development, waves are driven by glutamatergic rather than cholinergic synapses. Here, we asked how retinal activity evolves in the absence of cholinergic transmission by using a conditional mutant in which the gene encoding choline acetyltransferase (ChAT), the sole synthetic enzyme for acetylcholine (ACh), was deleted from large retinal regions. ChAT-negative regions lacked retinal waves for the first few days after birth, but by postnatal day 5 (P5), ACh-independent waves propagated across these regions. Pharmacological analysis of the waves in ChAT knock-out regions revealed a requirement for gap junctions but not glutamate, suggesting that patterned activity may have emerged via restoration of previous gap-junctional networks. Similarly, in P5 wild-type retinas, spontaneous activity recovered after a few hours in nicotinic receptor antagonists, often as local patches of coactive cells but not waves. The rapid recovery of rhythmic spontaneous activity in the presence of cholinergic antagonists and the eventual emergence of waves in ChAT knock-out regions suggest that homeostatic mechanisms regulate retinal output during development.

Characterization of Retrokeratoprosthetic Membranes in the Boston Type 1 Keratoprosthesis

OBJECTIVE To evaluate retroprosthetic membranes that can occur in 25% to 65% of patients with the Boston type 1 keratoprosthesis (KPro). METHODS Two patients with Peter anomaly and 2 with neurotrophic scarred corneas underwent revisions of their type 1 KPros because of visually compromising retroprosthetic membranes. The excised membranes were studied by light microscopy with hematoxylin-eosin, periodic acid-Schiff, and toluidine blue stains. Immunohistochemical and transmission electron microscopic examination were also used. RESULTS Light microscopic examination revealed that the retro-KPro fibrous membranes originated from the hosts corneal stroma. These mildly to moderately vascularized membranes grew through gaps in the Descemet membrane to reach behind the KPro back plate and adhere to the anterior iris surface, which had undergone partial lysis. In 2 cases, the fibrous membranes merged at the pupil with matrical portions of metaplastic lens epithelium, forming a bilayered structure that crossed the optical axis. Retro-KPro membranes stained positively for α-smooth muscle actin but negatively for pancytokeratin. Electron microscopy confirmed the presence of actin filaments within myofibroblasts and small surviving clusters of metaplastic lens epithelial cells. CONCLUSIONS Stromal downgrowth, rather than epithelial downgrowth, was the major element of the retro-KPro membranes in this series. Metaplastic lens epithelium also contributed to opacification of the visual axis. Florid membranous inflammation was not a prominent finding and thus probably not a requisite stimulus for membrane development. Further advances in prosthetic design and newer antifibroproliferative agents may reduce membrane formation.

Pythium insidiosum keratitis in Israel

Purpose: To report with morphologic and phylogenetic speciation the first case from Israel of Pythium insidiosum keratitis associated with contact-lens wear. Methods: Case report and literature review. Results: A 21-year-old man with a history of contact-lens use and water exposure was hospitalized in Israel for a corneal ulcer. The ulcer progressed despite intensive antibiotics. He flew home to the United States for further care. Examination revealed a corneal ulcer with hypopyon. The infection progressed despite intensive medical therapy, and a therapeutic penetrating keratoplasty was performed. Histology and cornea cultures from the host cornea revealed sparsely septate, branching hyphae, consistent with P. insidiosum. DNA sequencing of the Pythium isolate supported the clinical history that the infection was acquired outside of the United States. Despite intensive medical therapy and a second corneal transplant, the ulcer progressed, ultimately requiring enucleation. Conclusion: This is the first reported case of culture-proven, contact lens-related Pythium keratitis originating from Israel. Pythium is a fungus-like, aquatic oomycete found in tropical climates. Human pythiosis is uncommon but associated with high morbidity. Case reports describe surgical cure of Pythium keratitis, but this case recurred despite two penetrating keratoplasties and maximal antifungal therapy. In cases of presumed fungal keratitis that do not respond to antifungals, the fungus should be sent for speciation because early surgical intervention is the only means to save the eye in ocular pythiosis.

Diffuse large B-cell lymphoma of the orbit: clinicopathologic, immunohistochemical, and prognostic features of 20 cases.

PURPOSE To evaluate a series of orbital diffuse large B-cell lymphomas (DLBCL) for prognostic features and therapeutic outcomes. DESIGN Retrospective multicenter case study of clinical and immunohistochemical features of 20 patients. METHODS Clinical, histopathologic, and immunohistochemical features were correlated with outcomes. Immunohistochemistry for biomarkers including Bcl-6, CD5, CD10, CD20, FOXP1, GCET1, and MUM1 was performed to differentiate between 2 major genetic subtypes of DLBCL: activated B-cell-like (ABC) and germinal center B-cell-like (GCB). RESULTS Sixteen patients presented with unilateral and 4 with bilateral tumors. Three had bony erosion of the orbit on imaging studies. Of 14 patients with detailed follow-ups, 3 had a prior or concurrent lymphomatous disease; 8 had stage I disease (limited to the orbit) at presentation; and 3 were newly diagnosed with systemic (stage IV) DLBCL. Localized disease was treated with combined systemic chemotherapy, including rituximab and radiation with no deaths to date; there was 1 death related to systemic DLBCL. Clinical staging was the best predictive method and no immunohistochemical feature or subcategory (ABC vs GCB) correlated with outcome. CONCLUSIONS Primary orbital DLBCL has a more favorable prognosis than systemic DLBCL and may arise from a preexistent hematolymphomatous neoplasm (4 out of 20 cases). In our series, orbital DLBCL had a 57% likelihood of being restricted to the ocular adnexa. Clinical staging was more helpful in predicting outcome than any single immunohistopathologic feature or combination of biomarkers. Orbital radiation of 30 gray in conjunction with systemic chemotherapy with rituximab can achieve disease-specific survival approaching 100% in purely localized cases.

Orbital involvement in Bing-Neel syndrome.

Bing-Neel syndrome (BNS) is defined as intracranial involvement of Waldenström macroglobulinemia (WM). Few cases of orbital involvement have been reported. A 51-year-old man with a history of WM developed bilateral orbitopathy and optic neuropathy. Orbital biopsy, cerebrospinal fluid studies, and neuroimaging confirmed the diagnosis of BNS involving the orbital soft tissues, optic nerves, meninges, and cauda equina. The neuro-ophthalmic manifestations resolved after parenteral and intrathecal chemotherapy in addition to autologous stem cell transplantation. The rare neuro-ophthalmic manifestations of BNS may require a multifaceted approach to therapy.

IgG4-Positive Dacryoadenitis and Küttner Submandibular Sclerosing Inflammatory Tumor

Comment. C albicans is the most frequent cause of fungal keratitis in temperate regions and is an opportunistic organism that can complicate chronic keratopathy and corneal grafting. Persistent epithelial defects and suture-related problems, along with immunosuppression, have been found to be the major predisposing risk factors. Caspofungin is a first-in-class echinocandin with potent activity against Candida and Aspergillus, the dominant human fungal pathogens. In contrast to all other antifungal drugs, echinocandins have a selective action on a target present only in fungal cell walls (not in mammalian cells): they inhibit the synthesis of an essential component, (1,3)-D-glucan. Caspofungin is fungicidal in vivo and in vitro against all Candida species, including fluconazole-resistant strains. Its activity differs from that of the azole antifungal group, which is fungistatic. In our case, we think that voriconazole stopped progression of the infiltrate but did not kill the microorganism. The presence of the fungus after 1 month could be due to poor drug penetration, fungal resistance, or both. We therefore suggest that an ideal treatment protocol should include antifungal agents chosen on the basis of in vitro susceptibility of the fungus with a duration assessed by in vivo monitoring of fungal filaments or yeasts. To our knowledge, the topical ocular use of caspofungin has been reported in rabbits. There has been only 1 report of its use in humans, although it was in association with other antifungal drugs. In conclusion, topical caspofungin, 0.5%, is a new, promising option in the treatment of refractory fungalrelated corneal ulcers with no evidence of ocular toxic effects. However, future studies with larger samples are indicated to further evaluate its efficacy and tolerance.

Subtleties in the Histopathology of Giant Cell Arteritis

A temporal artery biopsy is typically obtained in cases of suspected giant cell arteritis (GCA). The differentiation between a “positive” versus a “negative” biopsy is sometimes not simple. Degrees of inflammation can vary from obvious, florid accumulations of giant cells to subtle pockets of non-granulomatous inflammation. Areas of normal pathology, or “skip lesions,” may be interspersed within inflamed sections of the artery, resulting in a false negative diagnosis. Other features of the biopsy, such as the state of the internal elastic lamina and inflammation of the adjacent, smaller vessels, must also be evaluated and considered as possible signs of disease. Finally, some biopsies have an intermediate histopathologic appearance with mild inflammation and scarring, which may indicate prior treatment with corticosteroids, or “healed” arteritis. Careful examination and clinical correlation is therefore essential to evaluate for these subtleties, which can affect the final diagnosis.

Pigmented apocrine hidrocystoma of the caruncle.

Purpose: To describe the clinical and immunopathologic features of the first convincing apocrine hidrocystoma of the caruncle that happened to be pigmented. Methods: Case report with clinical evaluation, ultrasound biomicroscopy, and paraffin-embedded tissue stained with hematoxylin and eosin, periodic acid-Schiff, and immunohistochemical markers. Results: Ultrasound biomicroscopy identified a cyst occurring in a 56-year-old white woman. Histopathologically, it was lined by an inner eosinophilic columnar epithelium that was gross cystic disease fluid protein-15-positive and evinced apical decapitation secretion and periodic acid-Schiff-positive, diastase-resistant cytoplasmic granules. Fontana- and MART-1-positive melanin granules were demonstrated. There was an outer layer of smooth muscle actin-positive myoepithelial cells. An adjacent apocrine gland was discovered with identical staining characteristics but without melanin granules. Conclusions: Classical apocrine gland cysts can exceptionally develop in the caruncle. The light dispersion of cytoplasmic melanin found in the lining cells may have contributed to the cysts clinical pigmentation, which is otherwise generally regarded as the result of the Tyndall effect, wherein blue wavelengths of light reflect from a colloidal solution of lipofuscin and cell debris.

Blue Nevus of the Tarsus as the Predominant Component of a Combined Nevus of the Eyelid

A 40-year-old man presented with a pigmented lesion of the palpebral conjunctiva and margin of the right lower eyelid. Because of suspicion of melanoma, the lesion was resected. Microscopic examination revealed 2 distinct components: a dominant blue nevus in the tarsus consisting of bland pigmented spindle and epithelioid cells that dissected among the orbicularis muscle fibers and meibomian glands, and a small subepithelial nevomelanocytic component with no overlying junctional activity. The diagnosis of a combined nevus was supported by minimal Ki-67 nuclear immunoreactivity. While the current lesion was proved to be an atypical nevus, all palpebral pigmented lesions should be routinely excised because many are melanomas.