Correlation of AMPA receptor subunit composition with synaptic input in the mammalian cochlear nuclei.

The composition of AMPA receptors in patches excised from somata and dendrites of six cell types in the mammalian cochlear nuclei was probed and compared electrophysiologically and pharmacologically with the rapid application of glutamate. Cells excited predominantly by auditory nerve fibers had AMPA receptors with exceptionally rapid gating (submillisecond deactivation and desensitization time constants). The nonlinear current-voltage relationship in the presence of spermine showed that few of these receptors had GluR2 subunits, and the insensitivity of desensitization to cyclothiazide indicated that they contained mostly flop splice variants. At synapses made by parallel fibers, AMPA receptors were slowly gating (time constants of deactivation and desensitization >1 msec) and contained higher levels of GluR2 and flip isoforms. However, receptors at auditory nerve synapses on cells that also receive parallel fiber input, the fusiform cells, had intermediate properties with respect to kinetics and contained GluR2 and flip isoforms. Given the diverse biophysical properties, patterns of innervation, patterns of electrical activity, and targets of each cell type in vivo, these data indicate that the kinetics and permeation properties of AMPA receptors are linked to factors associated with synaptic connectivity.

Detection of synchrony in the activity of auditory nerve fibers by octopus cells of the mammalian cochlear nucleus.

The anatomical and biophysical specializations of octopus cells allow them to detect the coincident firing of groups of auditory nerve fibers and to convey the precise timing of that coincidence to their targets. Octopus cells occupy a sharply defined region of the most caudal and dorsal part of the mammalian ventral cochlear nucleus. The dendrites of octopus cells cross the bundle of auditory nerve fibers just proximal to where the fibers leave the ventral and enter the dorsal cochlear nucleus, each octopus cell spanning about one-third of the tonotopic array. Octopus cells are excited by auditory nerve fibers through the activation of rapid, calcium-permeable, alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate receptors. Synaptic responses are shaped by the unusual biophysical characteristics of octopus cells. Octopus cells have very low input resistances (about 7 M Omega), and short time constants (about 200 microsec) as a consequence of the activation at rest of a hyperpolarization-activated mixed-cation conductance and a low-threshold, depolarization-activated potassium conductance. The low input resistance causes rapid synaptic currents to generate rapid and small synaptic potentials. Summation of small synaptic potentials from many fibers is required to bring an octopus cell to threshold. Not only does the low input resistance make individual excitatory postsynaptic potentials brief so that they must be generated within 1 msec to sum but also the voltage-sensitive conductances of octopus cells prevent firing if the activation of auditory nerve inputs is not sufficiently synchronous and depolarization is not sufficiently rapid. In vivo in cats, octopus cells can fire rapidly and respond with exceptionally well-timed action potentials to periodic, broadband sounds such as clicks. Thus both the anatomical specializations and the biophysical specializations make octopus cells detectors of the coincident firing of their auditory nerve fiber inputs.

Time course and permeation of synaptic AMPA receptors in cochlear nuclear neurons correlate with input.

AMPA receptors mediate rapid glutamatergic synaptic transmission. In the mammalian cochlear nuclei, neurons receive excitatory input from either auditory nerve fibers, parallel fibers, or both fiber systems. The functional correlates of differences in the source of input were examined by recording AMPA receptor-mediated, miniature EPSCs (mEPSCs) in whole-cell voltage-clamp mode from identified neurons. Bushy, octopus, and T-stellate cells of the ventral cochlear nucleus (VCN) and tuberculoventral cells of the dorsal cochlear nucleus (DCN) receive most of their excitatory input from the auditory nerve; fusiform cells receive excitatory inputs from both the auditory nerve and parallel fibers; cartwheel cells receive excitatory input from parallel fibers alone. mEPSCs from bushy, octopus, T-stellate, and tuberculoventral cells had significantly faster decay time constants (0.35-0.40 msec) than did those from fusiform and cartwheel cells (1.32-1.79 msec). Some fusiform cells had two populations of mEPSCs with distinct time courses. mEPSCs in cells with auditory nerve input alone were inhibited by philanthotoxin, a blocker of calcium-permeable AMPA receptors, whereas mEPSCs in cells with parallel fiber input were not. Thus AMPA receptors postsynaptic to the auditory nerve differ from those postsynaptic to parallel fibers both in channel-gating kinetics and in their permeability to calcium. These results confirm the conclusion that synaptic AMPA receptors are specialized according to the source of input (Hunter et al., 1993; Rubio and Wenthold, 1997; Wang et al., 1998).

Golgi cells in the superficial granule cell domain overlying the ventral cochlear nucleus: morphology and electrophysiology in slices.

Golgi cells are poised to integrate multimodal influences by participating in circuits involving granule cells in the cochlear nuclei. To understand their physiological role, intracellular recordings were made from anatomically identified Golgi cells in slices of the cochlear nuclei from mice. Cell bodies, dendrites, and terminals for all seven labeled cells were restricted to the narrow plane of the superficial granule cell domain over the ventral cochlear nucleus. The axonal arborization was the most striking feature of all Golgi cells; a dense plexus of terminals covered an area 200-400 microm in diameter in the vicinity of the cell body and dendrites. Axonal beads often surrounded granule cell bodies, indicating that granule cells are probable targets. Cells had input resistances up to 130 M omega and fired regular, overshooting action potentials. Golgi cells probably receive auditory nerve input, because shocks to the cut end of the auditory nerve excited Golgi cells with excitatory postsynaptic potentials (EPSPs). The latency of EPSPs shortened to a minimum and the amplitude of EPSPs grew in several steps as the strength of shocks was increased. The minimum latency of EPSPs in Golgi cells was on average 1.3 milliseconds, 0.6 milliseconds longer than the minimum latencies of EPSPs in nearby octopus and T stellate cells. The long latency raises the possibility that Golgi cells receive input from slowly conducting, unmyelinated auditory nerve fibers. Golgi cells are also excited by interneurons with N-methyl-D-aspartate receptors, probably granule cells, because repetitive shocks and single shocks in the absence of extracellular Mg2+ evoked late EPSPs that were reversibly blocked by DL-2-amino-5-phosphono-valeric acid.

Comparative ureteral microanatomy.

Several animals have been used as models for experimental upper urinary tract surgery. Although minipigs have recently gained favor because their upper urinary tract is grossly similar to that of humans, the ureteral microanatomic features of the two species have not been compared. We evaluated the histologic features of human and animal ureters in order to provide addition information regarding experimental models for upper urinary tract surgery. Cross-sections of upper, middle, and lower ureteral segments from human autopsy specimens and animals (farm pig, minipig, dog, sheep, rabbit, and rat) were evaluated after preparation with Masson's trichrome stain. Morphology, computer-assisted morphometry, linear micrometry, and point-count estimation of smooth muscle/connective tissue ratios were assessed. The epithelium was remarkably consistent across all species, both morphologically and in terms of absolute size. The lamina propria was highly vascularized in all species, but the compactness varied with the overall ureteral dimensions. In the muscularis, the relative size and the percentage composed of smooth muscle differed widely among species, as did the ratio of muscularis to lamina propria cross-sectional surface area. Among the species evaluated, the sheep, farm pig, and minipig ureters were the best histologic approximations of the human ureter. Given the difficulties with manipulation of the small sheep ureter, the minipig and farm pig models appear to be optimal for experimental research on upper urinary tract surgery by an endourologic approach.

The unintubated ureterotomy endourologically revisited.

PURPOSE: The duration of stenting after endoureterotomy is a controversial issue. An even more basic question, however, is whether stent placement is needed at all. We performed a porcine study to address this question. METHODS: A unilateral midureteral stricture was created in 20 minipigs. Six weeks later, 15 pigs underwent endoureterotomy. In 10 animals, a 7F ureteral stent was placed for 1 week. Five pigs remained unstented. Three months later, all 20 ureters were studied radiographically and were harvested. RESULTS: Three of 5 control animals and 2 of 10 stented animals has strictures. In the unstented study group, all 5 animals had a nonobstructing dilation at the site of endoureterotomy. This was corroborated by a normal furosemide washout renal scan in all 3 animals so tested. CONCLUSIONS: The result of unstented endoureterotomy in the porcine model suggests that, after endoureterotomy of secondary midureteral strictures, stenting may be unnecessary.

Comparison of acucise endopyelotomy and endoballoon rupture for management of secondary proximal ureteral stricture in the porcine model.

At this point in time, endopyelotomy is first-line therapy for both primary and secondary adult ureteropelvic junction obstruction (UPJO) in many medical centers. However, the potential, albeit small, for significant bleeding with any endoincision of the UPJ has sparked interest in the simple, less morbid technique of endoballoon rupture. To date, no comparative data are available on the effectiveness of these two techniques. Thirty female minipigs were randomized to cutting balloon (Acucise) endopyelotomy (AEP) (N = 13), endoballoon rupture (EBR) (N = 13), or a control arm (N = 4). Following baseline retrograde pyelogram (RPG) and diuretic renogram (DRG), a secondary proximal ureteral stricture was created by laparoscopic ligation of the UPJ. After 8 weeks, AEP or EBR was performed in each of the study pigs. In 16 pigs (8 AEP, 8 EBR), a 7F 22-cm ureteral stent was placed (chronic arm). After 6 weeks, the stent was removed, and a second RPG and DRG were performed. Three months post-treatment, after RPG and DRG, the renal units were harvested, and histologic sections of the affected UPJ, contralateral normal ureter, and ipsilateral kidney were examined. Ten pigs (5 AEP, 5 EBR) underwent harvest immediately after treatment (acute arm). The four control animals remained untreated. At 8 weeks, all minipigs had obstructive findings on RPG and DRG. All UPJs could be treated but one, which had an impassable stricture; there were no perioperative complications. In the acute arm, all UPJs were patent. All five AEP ureters had evidence of an uneven cut and cautery effect. Of the EBR ureters, two had smooth tears and three had ragged tears, and none had evidence of cautery effect. In the chronic arm, 3 months after either AEP or EBR, all minipigs had a patent UPJ, yet only 5 of 16 had an improved 1 1/2 by DRG. Histologic sections of the affected UPJs from 20 minipigs in the chronic arm (8 AEP, 8 EBR, 4 controls) were indistinguishable among the three groups; each revealed significant periureteral fibrosis and chronic inflammation with a mainly unremarkable muscular layer. However, histologic sections of 25 treated kidneys, including both acute and chronic animals (13 AEP, 12 EBR), revealed endstage renal disease (N = 10), chronic inflammatory changes (N = 7), or normal tissue (N = 8). Again, there was no trend favoring either AEP and EBR. In summary, in this laboratory study, we could detect no difference in outcome between an incisional endopyelotomy and an endoballoon rupture for treating secondary UPJO.

Laparoscopic bladder seromyotomy: laboratory experience.

Twelve female microminipigs with normal bladder capacity underwent laparoscopic bladder seromyotomy. Three other female microminipigs were added to the study as controls. Urodynamic studies and cystograms were performed preoperatively and at 2 and 4 months after a laparoscopic bladder seromyotomy. Histologic studies of the treated bladders were performed when the animals were euthanized at 4 months. The operation was completed in all 12 test animals. There was no significant difference in bladder capacity or leak-point pressure between the 12 seromyotomy and the 3 control bladders at any time point. Histologically, over the seromyotomy site, the urothelium remained intact; however, the muscularis was attenuated, and a thin layer of fibrotic tissue replaced the normal serosal covering of the bladder. Laparoscopic bladder seromyotomy can be performed reliably in the porcine animal model. Not surprisingly, no significant changes in bladder capacity or bladder compliance were seen in our study of normal bladders. An animal model of a neuropathic contracted bladder is needed to assess more accurately the physiological impact of laparoscopic seromyotomy.

Extracorporeal shock-wave lithotripsy of middle ureteral stones: are ureteral stents necessary?

OBJECTIVES: To ascertain whether insertion of a ureteral stent improves the outcome of middle ureteral (overlying the pelvic bone) stones treated with extracorporeal shock-wave lithotripsy (ESWL). METHODS: Thirty-three patients with middle ureteral stones were treated with ESWL at our institution between October 1991 and October 1994. Twenty-six patients were available for follow-up; 14 patients were treated with stent bypass, 8 were treated in situ, and 4 patients were treated after percutaneous nephrostomy (PCN). All patients were treated initially on an unmodified Dornier HM-3, and all but 4 patients were treated in the prone position on a modified Stryker frame. Follow-up consisted of a plain abdominal radiograph, intravenous urogram, occasionally a retrograde urogram, and a telephone interview. RESULTS: The overall stone-free rate for ESWL alone was 73%, and the efficiency quotient was 69. The stone-free rates after a single treatment for the stent bypass, in situ, and PCN groups were 71%, 63%, and 75%, respectively. Overall, 4% of patients required retreatment, 19% of patients required an auxiliary procedure, and 8% of the patients required hospital or emergency room admissions for renal colic. For stones 10 mm or greater (9), stone-free rates after one treatment for the stent bypass, in situ, and PCN groups were 33%, 33%, and 67%, respectively; for stones less than 10 mm (17), success rates were 82%, 80%, and 100%, respectively. CONCLUSIONS: Pretreatment stinting provides no advantage over in situ ESWL for middle ureteral calculi (Fisher's exact test, P = 1.0). ESWL is a reasonable initial therapy for middle ureteral stones less than 10 mm.

Comparison of newer laparoscopic port closure techniques in the porcine model.

Obtaining a precise fascial closure of larger laparoscopic port sites presents an ongoing challenge for even the most experienced laparoscopic surgeons. To this end, we evaluated five new methods for closing laparoscopic port sites and compared these with the standard technique of a hand-sutured repair using a 0 absorbable suture in the pig. The new devices studied were the Carter-Thomason Needle-Point Suture Passer (CTNSP), the eXit Disposable Puncture Closure (EDPC) device, the Maciol suture needle (MSN) set, the Endoclose suture carrier, and a nonpropreitary technique using a long 14-gauge Angiocatheter and a looped Prolene suture. Unlike the typical clinical situation, the fascia of the pig lies extremely close to the skin; hence, the sutured closure could be routinely performed most quickly of all. Among the newer techniques, the CTNSP provided the quickest closure. The Endoclose, MSN, and 14-gauge Angiocatheter techniques required the use of an auxiliary grasper, which appeared to slow the closure. Only the EDPC provided an airtight closure in 100% of cases. Of the new techniques, the 14-gauge Angiocatheter technique was the least expensive ($3). The efficacy of these techniques in the clinical realm needs to be assessed by a randomized trial.

Laparoscopic pneumodissection: a unique means of tissue dissection.

PURPOSE: To study the acute and chronic tissue effects of using high pressure CO2 to dissect tissues during laparoscopic procedures. MATERIALS AND METHODS: A 5-mm. laparoscopic pneumodissector, capable of delivering brief bursts of high pressure CO2 (20 to 100 psi), was discharged on a variety of porcine tissues including the renal hilum. Acute and chronic histological sections, intraoperative blood gases and perioperative renal scans were obtained. RESULTS: The only tissue with significant acute damage was the spleen. Acute changes in other tissues were minimal; long-term studies revealed no discernible damage to these same tissues. Intraoperative blood gases revealed no significant changes in the serum pCO2 or pH. Likewise, there was no renal damage discernible by renal scans. CONCLUSIONS: Pneumodissection at 50 psi is a safe method for dissecting the porcine kidney and the vessels of the renal hilum. Clinical studies are pending.

Evaluation of optimal stent size after endourologic incision of ureteral strictures.

The stent size necessary to promote optimal ureteral healing after an endoureterotomy is not known. We compared healing of an endoureterotomy over a 7F indwelling ureteral stent with healing over a 14F endopyelotomy stent. A midureteral stricture was created in each of 25 anesthetized female minipigs using an electrified stone basket passed retrograde. Six weeks later, the stricture was incised with a 24F cutting balloon device. Twenty pigs were randomized to receive a 7F or a 14F stent; four control pigs received neither incisions nor stents. At 1 week, a radiograph was performed to confirm proper stent position, and the stents were removed. At 3 months, a retrograde ureterogram was performed, and the ureters were examined grossly and harvested for histologic studies. Two of the ten pigs in each study group developed a recurrent stricture. The use of a 14F stent provided no advantage over the use of a smaller, more easily positioned 7F stent.

Comparison of retrograde endo-pyelotomy and endo-balloon rupture of the ureteropelvic junction in a porcine model.

Excellent results and durable success have been achieved with antegrade and retrograde endo-pyelotomy for treating primary and secondary ureteropelvic junction obstruction. Recently, a 30F dilating balloon was used to rupture the ureteropelvic junction (ENDOBRST) with encouraging results. While balloon distention of the ureteropelvic junction is a technically simpler procedure than endo-pyelotomy, clinical and laboratory data comparing the 2 methods are lacking. In an acute and chronic animal study we compared endo-pyelotomy via a ureteral cutting balloon incision to balloon rupture (that is 30F) of the normal ureteropelvic junction in each of 20 female farm pigs. Eight pigs were harvested acutely after treatment, and a macroscopic and histological examination of the treated ureteropelvic junction was completed. In 11 chronic pigs after endo-pyelotomy a 7F double pigtail ureteral stent was placed bilaterally and then removed after 6 weeks. Evaluation in the chronic group consisted of a furosemide washout renogram and retrograde pyelogram immediately preoperatively and 6 weeks after stent removal. The animals were likewise harvested 6 weeks after stent removal. One control pig underwent passage of the balloon cutting catheter and balloon dilating catheters without activation or dilation, respectively. Ureteral stents were placed bilaterally for 6 weeks and the pig was otherwise treated similarly to the other chronic study animals. In the acute group all ureters after endo-pyelotomy demonstrated retroperitoneal extravasation of contrast material. At harvest the ureters had been cleanly incised along a length of 3 to 4 cm. through the adventitial layer. In contrast, the balloon treated ureters showed free retroperitoneal extravasation in only half of the animals. Among the balloon treated ureters 7 of 8 had a linear tear of varying length (1 to 5 cm.) involving all but a thin adventitial layer of tissue. Histologically, the endo-pyelotomy ureters demonstrated a clean, linear transmural incision with virtually no destruction of surrounding tissue in 6 cases. In the remaining 2 cases an incision into but not completely through the muscular layer was observed. The balloon treated ureters showed a perforation through the muscular wall in 7 cases. However, periureteral hemorrhage and urothelial loss were common findings. In the chronic group infection and continued urine extravasation from the endo-pyelotomy site resulted in a 45% mortality rate. Of the surviving 6 pigs 83% of the balloon treated and 67% of the endo-pyelotomy pigs had a patent ureteropelvic junction by retrograde pyelogram and renogram. Histologically, the 2 sides were indistinguishable, with both showing mild fibrosis and chronic inflammation.(ABSTRACT TRUNCATED AT 400 WORDS)

Analyzing alertness among people with profound multiple disabilities: implications for provision of training.

We investigated whether variations in alertness among individuals with profound multiple disabilities affected subsequent responsiveness to training programs. Three experiments were conducted involving 5 people. In Experiment 1, results indicated that alertness of 3 individuals increased with provision of skill-acquisition training programs. Results also indicated no predictive relationship between alertness levels prior to training and subsequent responsiveness to training. Experiment 2 replicated results of Experiment 1 with 2 participants from Experiment 1 and an additional participant. Results of Experiment 2 also indicated that the increased alertness levels accompanying provision of training were not a function of the participants simply being in an inactive environment prior to training. Results of Experiment 3 indicated that alertness of an additional participant increased through provision of another training intervention, involving a systematic preference assessment. Responses during the preference assessment appeared to be unrelated to previously existing alertness levels. These findings suggest the need for caution when considering the withholding of scheduled training because an individual appears to be nonalert.

A comprehensive evaluation of reinforcer identification processes for persons with profound multiple handicaps.

We evaluated comprehensively a preference assessment for identifying reinforcers for persons with profound multiple handicaps. Four experiments were conducted involving 18 individuals. Results of Experiment 1 replicated previous findings in that the assessment identified student preferences for respective stimuli, and caregiver opinion of preferences did not coincide with the systematic assessment. Results of Experiment 2 indicated highly preferred stimuli were likely to function as reinforcers in training programs, whereas stimuli not highly preferred did not function as reinforcers. Results of Experiment 3 suggested the 12 stimuli used in the assessment represented a comprehensive stimulus set for identifying preferences, although the utility of the set sometimes could be enhanced by caregiver opinion. Results of Experiment 4 indicated the assessment identified preferences likely to be maintained over time. Overall, results are discussed in terms of identifying limits and alternatives to a behavioral teaching technology when applied to persons with profound multiple handicaps.

Increasing habilitative services for persons with profound handicaps: an application of structural analysis to staff management.

We evaluated a structural analysis methodology for enhancing the utility of a staff management program. In Experiment 1, a structural analysis of direct-care staff behavior in a mental retardation facility revealed differences in work patterns over time. Specific times were identified when few basic care duties were necessary and staff engaged in nonwork activity. In Experiment 2, a management program was implemented to increase staff members' training activities during periods identified through the structural analysis. The program was accompanied by increases in training activities and decreases in nonwork behavior. The improvements were maintained during a 43-week period while the most labor-intensive component of the program was withdrawn. Staff acceptability measures indicated a positive response to the management intervention, although responses varied across components within the multifaceted program. The increased training was accompanied by beneficial changes among clients with profound handicaps. Results are discussed regarding practical considerations for improving staff performance and for adopting innovations resulting from applied research.

Identifying reinforcers for persons with profound handicaps: staff opinion versus systematic assessment of preferences.

We evaluated a systematic means of determining stimulus preferences among seven profoundly handicapped persons. Preferences were determined by observing student approach responses to individual stimuli. Results indicated that there were differential stimulus preferences across the multiply handicapped participants. However, results of the systematic assessment did not coincide with the results of a more traditional, caregiver-opinion method of assessing student preferences. A second experiment was then conducted with five participants to evaluate whether stimuli that were assessed to consistently represent preferences would function as reinforcers in skill training programs. Results indicated that stimuli that were systematically assessed to represent student preferences typically functioned as reinforcers when applied contingently. However, preferred stimuli as reflected by caregiver opinion did not function as reinforcers unless those stimuli were also preferred on the systematic assessment. Results are discussed in terms of assisting profoundly handicapped persons by (a) improving the effectiveness of training programs by increasing the likelihood of using stimuli that have reinforcing value and (b) increasing the overall quality of life by providing preferred stimuli in the routine living environment.

Quantitative molecular hybridization with unfractionated, solubilized cells using RNA probes and polyacrylamide gel electrophoresis.

Molecular hybridization with RNA probes was performed on unfractionated cells solubilized in guanidine thiocyanate solutions. Unhybridized probe was digested with ribonuclease, and protected probe fragments were resolved by polyacrylamide gel electrophoresis (PAGE). Since the same medium was used both for solubilization of the cells and as the hybridization buffer, RNA purification was not required and the analysis of large numbers of samples was facilitated. Using this method, specificity is superior to dot blot analysis because the size of hybridized fragments is determined and the signal of the probe hybridized to target RNA is separated from the background by PAGE.

Mouse lymphotoxin and tumor necrosis factor: structural analysis of the cloned genes, physical linkage, and chromosomal position.

Lymphotoxin (LT) and tumor necrosis factor (TNF) are cytotoxic and immunoregulatory lymphokines which have similar activities but are produced by different cell types. We have cloned the murine LT and TNF genes from a lambda:mouse DNA recombinant library, using as probes synthetic oligonucleotides defined by portions of the human LT or TNF cDNA sequences. Analysis of the genomic clones indicates that the LT and TNF genes are physically linked, i.e., approximately 1.2 kb separates the 3' end of LT from the 5' end of TNF genes. By using, first, a series of recombinant inbred lines, and second, a series of H-2-recombinant congenic strains, we determined that the LT/TNF gene cluster lies on chromosome 17, closely linked to the H-2D end of the murine H-2 complex. Comparison of the primary sequence of murine and human LT revealed that the intron-exon structure of murine LT is similar in these two species. Comparison of the predicted amino acid sequences of murine and human LT indicates that the proteins are about 72% homologous with much greater sequence conservation in regions encoding the COOH-terminal portion. Comparison of the 5' flanking sequence of LT to a number of genes that are specifically expressed in activated T cells reveals a number of conserved sequences that may play a role in control of these genes.

Griffiths Scales of Mental Development and different users.

In a study of the inter-rater reliability of the Griffiths Mental Development Scales, the Eye-Hand Coordination and the Performance and Practical Reasoning Scales were found to show greater consistency between raters than the Locomotor, Personal-Social scale and the Hearing-Speech Scale. The latter scales were more sensitive to interpretation by different assessors, even when all viewed the same videotaped performance. No significant discrepancies were found between raters with different backgrounds and experience.