Secondary Membranes Research Articles

Simultaneous Bilateral Cataract Surgery in Premature Babies With and Without Retinopathy of Prematurity

Background:Four premature babies (eight eyes) undergoing simultaneous bilateral cataract surgery are presented and discussed.Methods:All four babies underwent simultaneous bilateral cataract surgery. Three babies (six eyes) had primary implantation of posterior chamber intraocular lenses (IOLs) and one baby (two eyes) had primary lensectomies with secondary visual correction with contact lenses. Results: In all eight eyes, there was no endophthalmitis and no spontaneous choroidal hemorrhages. All eyes experienced large myopic shifts, as high as –15.00 D. All six eyes with IOLs required secondary membranectomies, which did not reoccur. Case 4 had Lowe’s syndrome, was bilaterally aphakic post-op, and subsequently developed glaucoma requiring bilateral glaucoma surgery. Conclusions: Simultaneous bilateral cataract surgery in severely premature babies can be successful in restoring vision over the long term. Strategies to successfully deal with the timing of surgery, IOLs, secondary membranes, secondary glaucoma, appropriate IOL powers, and IOL formulas is discussed. Successful long-term successful visual outcomes are now possible in this complex group of premature babies.

Gene Expression ofIGF1,IGF1R, andIGFBP3in Epiretinal Membranes of Patients with Proliferative Diabetic Retinopathy: Preliminary Study

The molecular mechanism formation of secondary epiretinal membranes (ERMs) after proliferative diabetic retinopathy (PDR) or primary idiopathic ERMs is still poorly understood. Therefore, the present study focused on the assessment of IGF1, IGF1R, and IGFBP3 mRNA levels in ERMs and PBMCs from patients with PDR. The examined group comprised 6 patients with secondary ERMs after PDR and the control group consisted of 11 patients with idiopathic ERMs. Quantification of IGF1, IGF1R, and IGFBP3 mRNAs was performed by real-time QRT-PCR technique. In ERMs, IGF1 and IGF1R mRNA levels were significantly higher in patients with diabetes compared to control subjects. In PBMCs, there were no statistically significant differences of IGF1, IGF1R, and IGFBP3 expression between diabetic and nondiabetic patients. In conclusion, our study indicated IGF1 and IGF1R differential expression in ERMs, but not in PBMCs, of diabetic and nondiabetic patients, suggesting that these factors can be involved in the pathogenesis or progression of proliferative vitreoretinal disorders. This trial is registered with NCT00841334.

IDIOPATHIC AND SECONDARY EPIRETINAL MEMBRANES

The purpose of this study was to define the morphologic differences in idiopathic and secondary epiretinal membranes (ERMs) using a time domain optical coherence tomography. The medical records and optical coherence tomography images of 293 eyes of 236 consecutive patients diagnosed to have ERM were evaluated retrospectively. Demographic features, best-corrected visual acuities, central macular thickness, membrane attachment patterns, macular changes, and the presence of posterior retinal detachment at the time of diagnosis were compared between the groups (idiopathic group, n = 125 eyes; secondary group, n = 168 eyes). The mean age was higher among idiopathic ERMs (67 ± 9 vs. 62 ± 10 years, P = 0.001). In the secondary group, best-corrected visual acuity was estimated to be worse (P < 0.001) and central macular thickness higher (P = 0.02) than the idiopathic group. In both groups, ERMs were mostly diagnosed at Grade 1 level. Diffuse attachment was more common in both of the groups. Among macular changes, only cystoid macular degeneration differed significantly between the groups (P = 0.0001). Idiopathic and secondary ERMs were estimated to differ significantly at the time of diagnosis in terms of age, visual acuities, macular thickness, presence of cystoid macular degeneration, and posterior vitreous detachment. These parameters may assist the retinal surgeon in the treatment process of ERM.

TMP revisited: the importance of plasma colloid osmotic pressure in high-flux dialysers

With high-flux dialysers and volume-controlled machines, transmembrane pressure (TMP) is no longer used to prescribe and control the adjustment of patient volume but still used to monitor filter function and membrane integrity. A drop in TMP at given operation characteristics indicates a leak or even a rupture in the membrane, while an increase in TMP indicates a loss in hydraulic permeability, most likely because of gradual clogging of the filter by formation of secondary membranes or obstruction of individual capillary fibers. Monitoring dialyser function is of special interest in different forms of haemofiltration and haemodiafiltration, where ultrafiltration rates are allowed to reach very high values exposing all components of

EPIRETINAL MEMBRANES INDICATE A SEVERE PHENOTYPE OF NEUROFIBROMATOSIS TYPE 2

The purpose of this study was to describe a subset of severely affected patients with neurofibromatosis type 2 (NF2), multiple central nervous system tumors, and characteristic retinal lesions. This is a retrospective observational case series of 4 patients with NF2. The time domain-optical coherence tomography findings of three patients have previously been described in another series. Ophthalmic signs were identified at a mean age of 6 years, and NF2 was diagnosed at a mean age of 11 years. Patients presented with diminished visual acuity in one or both eyes and epiretinal membranes in the absence of posterior vitreous detachment. The biomicroscopic and optical coherence tomography features were distinct from secondary epiretinal membranes or combined hamartomas of the retina and retinal pigment epithelium and pathognomonic for NF2. The ophthalmic manifestations were recognized before neurologic signs and led to the diagnosis of NF2 in 3 of the 4 patients. Each patient had > or =2 central nervous system tumors at the time of diagnosis, and 3 of 4 eventually required neurosurgical interventions for symptomatic, compressive lesions at a mean age of 12 years. Recognition of epiretinal membranes with a characteristic optical coherence tomography appearance may permit early diagnosis in neurologically asymptomatic children with a severe phenotype of NF2.

Simultaneous Surgery in Bilateral Congenital Cataract

To assess the efficiency of simultaneous surgery in bilateral congenital cataract not only in patients with a high anesthesiologic risk profile. A retrospective study was carried out on 40 bilateral congenital cataract patients (80 eyes) who presented to the outpatient department of pediatric ophthalmology of the University Federico II of Naples in the period from 1990 to 2005. All patients had undergone cataract extraction from both eyes in a single surgical session. Visual rehabilitation was achieved in all patients by corneal lenses; successively 25 patients had a secondary intraocular lens implant in the posterior chamber after a period of 2.5-3 years. The mean age at cataract surgery was 7 months (1-17 months). Visual acuity (VA) was assessed in 52 eyes: 8 eyes (15%) presented VA <or=20/200, 21 eyes (41%) showed VA 20/200 < 20/40, 23 eyes (44%) presented VA >or=20/40. Ocular motility disorders were seen in 28 patients (70%). Eight patients had postoperative complications: the formation of secondary membranes in five patients, and secondary glaucoma in three patients. There were no cases of endophthalmitis. Simultaneous surgery in bilateral congenital cataract may be taken into consideration not only for patients with a high anesthesiologic risk profile.

Xylem – as well as phloem – sustains severe damage due to feeding by the Russian wheat aphid

Investigation of comparative effects of feeding damage by the Russian wheat aphid (RWA, biotype SA1, Diuraphis noxia Mordvilko) on leaf blades of susceptible and resistant wheat cultivars ( Triticum aestivum L. var Betta and Betta- Dn1 respectively) were carried out to establish the level of ultrastructural damage caused by this aphid and the possible limitation of damage induced which could be ascribed to the resistance gene Dn1 over the susceptible cultivar. Ultrastructurally, Betta- Dn1 sustained less damage to the vascular tissue as well as to the mesophyll during the experimental period. Both inter- and intracellular probes resulted in considerable saliva deposition as the aphids probed for suitable feeding sites. Salivary tracks were observed between and within mesophyll, bundle sheath cells as well as the vascular tissue, including the xylem. Disruption of organelles and cytoplasm resulted from cell probing and sheath deposition. Cell and organelle damage was more evident in the non-resistant Betta cultivar. The aphids probed for and fed from thin-walled sieve tubes preferentially. Few thick-walled sieve tubes showed evidence of either aphid probing or feeding-related damage. Saliva was deposited when the aphids probed inter- and intracellularly for feeding sites. The aphids appeared preferentially to probe for and feed from thin-walled sieve tubes, as few thick-walled sieve tubes showed evidence of damage. Vessels, apparently probed for water, contained watery saliva that encased the secondary walls and sealed pit membranes between probed vessels and xylem parenchyma. The xylem probed by the RWA was rendered non-functional, probably contributing to symptoms of leaf roll, chlorosis and necrosis, which were observed within two weeks of infestation in the susceptible Betta cultivar. This damage was limited in the resistant Betta- Dn1 cultivar during the same time frame.

Secondary Membranectomy in Pediatric Pseudophakia Through a Limbal Approach

Purpose: Secondary membranes are a common problem following pediatric cataract extraction. In many cases it is of sufficient density to cause obscuration of the visual axis and requires removal. While in adults this is usually accomplished as an in-office laser procedure, most pediatric patients must return to the operating room for surgical removal. This has been accomplished with intraoperative YAG laser, a pars plana/posterior approach, or a limbal approach. Method: We performed 25 secondary membranectomies using a limbal approach.

Modification of NF membrane properties by in situ redox initiated graft polymerization with hydrophilic monomers

Nanofiltration membranes used for treating wastewater streams can be modified to change their adsorption and solute rejection properties and their susceptibility to fouling. NF200 membranes modified by hydrophilic monomer such as PEGMA showed greatly reduced adsorption of certain organic compounds typical of paper mill streams. Further testing of these membranes and NF270 in flat sheet and spiral element form showed indications that the modification gave modest increases in resistance to fouling by acid clear filtrate from paper mills. In tests on tertiary effluent from wastewater treatment plants, modification of NF membranes (NF70, NF270) by HEMA lead to significantly reduced salt passage, especially of monovalent anions (chloride). The effects of modification on solute retention were much more pronounced on the tertiary effluent. These effects can be partly explained in terms of the relative role of ordered water and secondary membranes formed by deposits on the membrane.

Secondary potential in electrodialysis membranes and the effect on permselectivity

The most important factor in the electrodialysis (ED) process is the permselectivity of the ion exchange membranes, which permit not only the separation of cations and anions in a solution, but also the separation of ions with the same sign but different valences. In this work, the mechanism of the permselectivity has been studied through the measurement of the potentials at different planes of the membrane. The experimental results have shown that there was a secondary potential inside ion exchange membranes in an electrodialysis process. At the membrane side touched with dilute solution, this secondary potential enhanced the external electrical field, and thus speeded up the passage of the corresponding ions in the dilute solution through the membranes; at the membrane side touched with concentrated solution, the secondary potential was contrary to the external electrical field and thus counteracted it, which could be very helpful by preventing the ions in the concentrated solution from entering the membranes. Obviously, the existence of the secondary potential might play an important role in the permselectivity of ion exchange membranes in ED processes.

Secondary membranes for flux optimization in membrane filtration of biologic suspensions.

We employ in situ deposited secondary membranes of yeast (SMYs) to optimize permeate flux during microfiltration and ultrafiltration of protein solutions. The deposited secondary membrane was periodically removed by backflushing, and a new cake layer was deposited at the start of the next cycle. The effects of backflushing time, backflushing strength, wall shear rate, and amount of secondary membrane deposited on the permeate flux were examined. Secondary membranes were found to increase the permeate flux in microfiltration by severalfold. Protein transmission was also enhanced owing to the presence of the secondary membrane, and the amount of protein recovered was more than twice that obtained during filtration of protein-only solutions under otherwise identical conditions. In ultrafiltration, the flux enhancement owing to the secondary membrane was only 50% or less. In addition, the flux for ultrafiltration was relatively insensitive to changes in the concentration of yeast used during deposition of SMY and to the backflushing strength used to periodically remove the secondary membrane.

Complications in infants undergoing surgery for congenital cataract in the first 12 weeks of life: is early surgery better?

To determine if there is an association between the timing of surgical intervention for congenital cataract within the first 12 weeks of life and the prevalence of postoperative complications. We performed a retrospective review of records from 1990 to 2000 of infants who underwent surgery for congenital cataract within the first 12 weeks of life. Eighty eyes in 55 children were involved with a minimum follow up of 6 months. Bilateral cataracts were present in 25 and monocular cataracts in 30 infants. A limbal approach lensectomy-vitrectomy was performed in all infants. Children with aphakia were rehabilitated with contact lens or glasses. Operative and postoperative complications-including glaucoma, nystagmus, strabismus, retinal detachment, and posterior capsule opacification/secondary membranes-were recorded. Ocular and systemic associations were noted. Statistical analysis was carried out with classification and regression trees (CART). The mean age at the time of surgery was 31.5 +/- 23.3 days (median, 26.5; range, 2 to 84). Mean follow up from the time of surgery was 2.85 +/-1.9 years (median, 2; range, 0.5 to 8). Persistent fetal vasculature (persistent hyperplastic primary vitreous) was present in 14 eyes. One infant with bilateral persistent fetal vasculature had bilateral retinal dysplasia and was excluded from the analysis. Glaucoma developed in 12 infants (22%); nystagmus was present in 18 infants (33%); strabismus developed in 28 infants (52%); and secondary membranes developed in 7 eyes (13%). CART analysis suggests that glaucoma is more prevalent in infants when the surgery was performed between 13.5 and 43 days of life (CART = 0.370); nystagmus when surgery is performed between 48 and 84 days of life (CART = 0.500); strabismus when surgery is performed between 55.5 and 84 days of life (CART = 0.600); and secondary membranes when surgery is performed between 26.5 and 40 days of life (CART = 0.4). Our data suggest that the first 2 weeks of life comprise the most favorable time for decreasing postoperative complications resulting from surgical intervention for infants presenting with cataracts within the first 12 weeks of life.

Ultrastructural localization of flotillin-1 to cholesterol-rich membrane microdomains, rafts, in rat brain tissue

There is much interest in research on cholesterol-rich membrane microdomains, rafts, in the field of neurobiology. However, no one has shown the ultrastructure of rafts in tissues. We examined the ultrastructure of rafts in rat brain tissue by pre-embedding immunoelectron microscopy using flotillin-1 antibody, which is a biochemical marker of lipid rafts, and BCθ, which is nicked and biotinylated θ-toxin, and binds to membrane cholesterol of rafts. Flotillin-1- and BCθ-labeled areas were patchy and prominent on the plasma membranes of small processes and synapses in the neuropil. The size of flotillin-1 labeling was 40–200 nm. In addition, the membrane of lysosome and Golgi apparatus were frequently labeled for flotillin-1 with a patchy pattern. Flotillin-1 and BCθ were mostly colocalized in double immunolabeling on a part of the plasma membranes of small processes and secondary lysosome membranes. We first indicate that flotillin-1 localizes to BCθ-positive cholesterol-rich membrane microdomains in vivo, and that flotillin-1 and BCθ could be ultrastructural raft markers in neural tissue.

Prospective analysis of pediatric pseudophakia: Myopic shift and postoperative outcomes

Purpose: Limited data exist about long-term refractive changes in eyes of children with intraocular lens (IOL) implantation. Information of postoperative results should allow more accurate predictions for IOL power implantation in children. Data regarding IOL complications, including secondary membranes, myopic shift, stereopsis, and pseudophakic glaucoma should also be reported. Methods: In a prospective study, the refractive errors of all pediatric patients between 12 months and 18 years who had cataract surgery and IOL implantation were evaluated at 4 weeks, 3 months, 6 months, 1 year, and every 6 months thereafter. All patients were followed for a minimum of 3 years. Results: Fifty-two eyes of 42 patients met inclusion criteria. Forty-two eyes had developmental cataracts. There were 10 bilateral cases. Of the 52 eyes, 85% had 20/40 vision or better. Visual acuity of 20/30 or better was achieved in 95% of bilateral eyes. In unilateral cataracts, visual acuity was 20/50 or better in 74% of eyes. Mean follow-up time was 5.45 years with a range of 3 to 10.5 years. Mean follow-up by age group ranged between 4.38 and 6.35 years. Children operated on at 12 months to 2 years of age had a mean myopic shift of −5.96 D; children operated on at 3 and 4 years of age had a −3.66 D shift; children operated on at 5 and 6 years of age had a shift of −3.40 D; children operated on at 7 and 8 years of age had a shift of −2.03 D; children operated on at 9 and 10 years of age had a mean shift of −1.88 D; children operated on at 11 to 14 years of age had a shift of −0.97 D; children operated on at 15 to 18 years of age had −0.38 D shift. No cases of pediatric pseudophakic glaucoma were observed. Secondary membrane occurred in 72% of eyes when the capsule was left intact. The operated eye showed a greater mean myopic shift than the nonoperated eye. No statistically significant difference in refractive change was found comparing amblyopic to nonamblyopic eyes or traumatic to nontraumatic cataracts. Conclusions: The greatest rate of refractive growth or change occurred between 1 and 3 years of age. After age 3 years, the rate of refractive growth followed a more linear trend. Based on this study, we have provided a guide for selecting IOL power in pediatric cataract cases using current formulas with the understanding that new formulas will need to be devised to better predict IOL power in children. (J AAPOS 2002;6:277-82)

Elimination of levofloxacin in critically ill patients with renal failure: influence of continuous veno-venous hemofiltration.

Pharmacokinetic data on levofloxacin in critically ill patients are sparse and conflicting. Aim of the study was to assess the clearance of levofloxacin in critically ill patients treated with continuous veno-venous hemofiltration (CVVH). Pharmacokinetics of levofloxacin were studied in 11 critically ill patients. Four patients were treated with CVVH because of renal failure, 4 patients had moderately impaired renal function but were not on hemofiltration, and 3 patients had approximately normal renal function. Patients received 0.5 g levofloxacin infused over 0.5 hours. Plasma levels of levofloxacin were determined by HPLC and pharmacokinetic parameters were calculated using a non-compartmental model. Levofloxacin clearance in critically ill patients with approximately normal renal function was similar to that in healthy subjects. In critically ill patients with impaired renal function not on CVVH, mean half-life was prolonged by a factor of about 3 (20-25 hours). The mean residence time and the volume of distribution were also increased. In renal failure treated with CVVH, a wide variability in pharmacokinetics was seen. The half-life was about 30 hours and the mean levofloxacin clearance was raised by a factor of 2. The area under the concentration-time curve was reduced by hemofiltration, while the volume of distribution was increased. There was a positive correlation between blood flow through the hemofilter and levofloxacin clearance. Variable amounts of the drug were recovered from the hemofilter. Most plasma levels, however, were in the therapeutic range and drug accumulation to toxic plasma concentrations was not observed in renal failure patients undergoing CVVH and receiving single daily administration of 0.5 g of levofloxacin i.v. During CVVH using polysulfone membrane hemofilters, plasma concentrations of levofloxacin are not easily predictable. Levofloxacin clearance may be affected by binding to secondary membranes formed in hemofilters during CVVH and blood flow rates have a significant impact on the pharmacokinetics of levofloxacin.

Surgical management of congenital cataract associated with severe microphthalmos

Purpose To report the results of cataract surgery in children with severe microphthalmos and congenital cataract. Setting Seoul National University, Department of Ophthalmology, Seoul, Korea. Methods Retrospective studies were conducted by reviewing the charts of 20 eyes of 11 patients with severe microcornea and microphthalmos or severe microcornea with a corneal diameter smaller than 9.0 mm. The patient pool was divided according to surgical method, specifically by incision site: an anterior group that had corneal limbal incisions and a posterior group treated via pars plicata incisions. Preoperative examinations included bilaterality, corneal diameter, axial length, and eye and systemic abnormalities. Postoperative results and complications of the 2 operative methods were compared. Results All 11 patients had bilateral congenital cataract and microphthalmos. Except in 1 case, surgeries were done in patients ranging in age from 2 to 16 months. Mean follow-up was 2 years. Corneal diameter was smaller than 9.0 mm in all 20 eyes and smaller than 7.5 mm in 11 eyes. Ten eyes were categorized into the anterior group, and the other 10 eyes were placed in the posterior group. Postoperative complications included secondary membranes in 3 eyes in the posterior group and corneal opacity in 3 eyes in the anterior group. Conclusion The surgical management of children with congenital cataract and severe microphthalmos is recommended as a way to improve vision but must be performed carefully to avoid complications.

Cataract surgery in children with chronic uveitis

Objective To evaluate the visual outcome of cataract surgery in children’s eyes with chronic uveitis and the feasibility of intraocular lens (IOL) implantation in these cases. Design Retrospective noncomparative case series. Participants Seventeen children (20 eyes) with chronic uveitis, dense cataract, and a preoperative visual acuity of 6/120 or less with follow-up of 5 years after the initial cataract surgery. Methods In 10 eyes of 10 children (five with juvenile rheumatoid arthritis [JRA] and five with non-JRA-associated uveitis) with uniocular or markedly unequal binocular disease, surgery was carried out through the limbus and a posterior chamber IOL was implanted. In seven children (10 eyes), three with JRA and four with non-JRA-associated disease, a pars plana approach was used, and contact lenses or glasses (for the bilateral cases) were prescribed. Results The postoperative course and immediate restored visual acuities were similar whether an IOL was implanted or not. One month after the surgery, visual acuity improved in all operated eyes. After monocular surgery, in the younger children, contact lenses were poorly tolerated and their use discontinued. These aphakic eyes remained with low vision, developing strabismus on longer follow-up. Children with JRA-associated uveitis were younger, demonstrated an active intraocular inflammation for an extended period after surgery, and tended to have secondary membranes develop, necessitating a second surgical intervention. Five years after the initial surgery, only two of nine eyes (22%) in the JRA group (one aphakic of a bilaterally affected child and one pseudophakic in a child undergoing cataract surgery in one eye) retained a visual acuity of 6/9 and 6/6, respectively. In the other seven eyes, the visual acuity was 6/60 in one pseudophakic eye and 6/240 or less in six eyes (three aphakic and three pseudophakic). In children with non-JRA-associated uveitis, 6 (four aphakic in two patients bilaterally affected and two pseudophakic) of 11 eyes (54.5%) retained a vision of 6/12 or better. Conclusions Cataract surgery in children’s eyes with uveitis may be beneficial. IOL implantation seems preferable to correction with contact lenses in young children needing surgery in one eye. In children with JRA-associated uveitis, the final visual results remain guarded because of irreversible amblyopia and a more complicated postoperative course. For these cases, a modified management approach and a better surgical technique are needed.

Modeling of fouling reduction by secondary membranes

The external cake of rejected particles plays an important role in filtration processes, often influencing the permeation rates of all species. It may have higher resistance to the permeate flow than the clean membrane itself, but it may also act as a secondary or dynamic membrane which screens the primary membrane from the more strongly fouling species of smaller size. The screening process due to the presence of the secondary membrane is modeled for filtration of a bidisperse suspension of large and small particles. The large particles form an external cake on the surface of the primary membrane, which then acts as a deep-bed filter and captures some of the small particles and prevents them from fouling the primary membrane. The theory predicts an optimum concentration of the large particles which maximizes the permeate volume for a given duration of filtration. To test the model, cross-flow microfiltration experiments were done with yeast suspensions, protein solutions, and yeast–protein mixtures using cellulose acetate membranes. During filtration of yeast–protein mixtures, protein aggregates were captured or screened by the yeast cake, which reduced fouling of the primary membrane by the protein aggregates. Under optimum conditions, the presence of yeast increased the total permeate volume by two fold.

Effect of fractal flocculation behavior on fouling layer resistance during apple juice microfiltration

The colloidal material in juice obtained from Red Delicious apples held in cold storage for over 6 months was observed to spontaneously aggregate to form ordered flocculation patterns. These flocculation patterns could be altered by heat treatment and by gelatin or antioxidant addition. The specific resistance of the fouling layer produced by each juice during microfiltration could be qualitatively predicted by examining the structure of macroscopic flocculation patterns. Environmental scanning electron microscopy (ESEM) photos of the various hydrated fouling layers indicated that loose flocs appeared to compress and rearrange on the membrane surface, producing smooth, low porosity secondary membranes. Densely packed cross-linked aggregate networks also formed low porosity structures, once again creating a fouling layer with high resistance. The lowest resistance fouling layers were produced when the flocs were dense enough to resist compression but porous enough to provide pathways for permeate flow. Field emission scanning electron microscopy (FESEM) and ESEM images indicated that, in many cases, the spacial distribution of solids in the microscopic aggregates were reminiscent of those in the macroscopic flocs. Such scaling relationships are typical of fractal systems.

Polysialylated NCAM expression during motor axon outgrowth and myogenesis in the fetal rat.

Polysialylation of the neural cell adhesion molecule (NCAM) converts it into an anti-adhesive molecule, attenuating intercellular adhesion and repelling apposed membranes. Previous studies have demonstrated that interaxonal repulsion, or defasciculation, induced by polysialylated NCAM (PSA-NCAM) expressed along outgrowing chick motor axons promotes intramuscular branching and facilitates differential guidance of segregating axonal populations. In the present study, we have examined the expression of PSA-NCAM in a developing mammalian motor system during axonal outgrowth, separation of distinct axonal populations, and intramuscular branching. Furthermore, we provide the first clear demonstration of the spatiotemporal modulation of PSA-NCAM expression on myotubes during each stage of myogenesis. Immunohistochemical labelling was used to compare the spatiotemporal pattern of PSA-NCAM expression with those of total-NCAM, the cell adhesion molecule L1, and growth associated protein (GAP-43) during development of the phrenic nerve and diaphragm of fetal rats (embryonic days, E11-E19). During segregation of phrenic and brachial axonal populations at the brachial plexus (E12.5-E13), PSA-NCAM expression was restricted to phrenics, being absent from brachial motoneurons. Both populations labelled equivalently for NCAM, L1, and GAP-43. We postulate that PSA-NCAM may be a component of the molecular machinery that specifically guides phrenic motoneuron growth at the brachial plexus. During diaphragmatic morphogenesis, PSA-NCAM expression: (i) remained high within the phrenic nerve throughout intramuscular branching; (ii) was transiently up-regulated on myotubes during myotube separation associated with primary and secondary myogenesis; (iii) was restricted to those regions of primary and secondary myotube membranes, which were juxtaposed and about to separate. These data suggest a role for PSA-NCAM in the guidance of specific subsets of mammalian motoneurons and in intramuscular branching, and demonstrate an intimate correlation between PSA-NCAM expression and myotube separation.