Refinement of detecting atrial fibrillation in stroke patients: results from the TRACK-AF Study.

BACKGROUND AND PURPOSE: Detection of occult atrial fibrillation (AF) is crucial for optimal secondary prevention in stroke patients. The AF detection rate was determined by implantable cardiac monitor (ICM) and compared to the prediction rate of the probability of incident AF by software based analysis of a continuously monitored electrocardiogram at follow-up (stroke risk analysis, SRA); an optimized AF detection algorithm is proposed by combining both tools. METHODS: In a monocentric prospective study 105 out of 389 patients with cryptogenic stroke despite extensive diagnostic workup were investigated with two additional cardiac monitoring tools: (a) 20 months' monitoring by ICM and (b) SRA during hospitalization at the stroke unit. RESULTS: The detection rate of occult AF was 18% by ICM (n = 19) (range 6-575 days) and 62% (n = 65) had an increased risk for AF predicted by SRA. When comparing the predictive accuracy of SRA to ICM, the sensitivity was 95%, specificity 35%, positive predictive value 27% and negative predictive value 96%. In 18 patients with AF detected by ICM, SRA also showed a medium risk for AF. Only one patient with a very low risk predicted by SRA developed AF revealed by ICM after 417 days. CONCLUSIONS: A combination of SRA and ICM is a promising strategy to detect occult AF. SRA is reliable in predicting incident AF with a high negative predictive value. Thus, SRA may serve as a cost-effective pre-selection tool identifying patients at risk for AF who may benefit from further cardiac monitoring by ICM.

Computational models for detection of endocrinopathy in subfertile males.

The observation that men with sperm density greater than 10 million/ml had low probability of endocrinopathy led to a refinement in the evaluation of subfertility. Using statistical methods, we sought to provide a more accurate prediction of which patients have an endocrinopathy, and to report the outcome as the odds of having disease. In addition, by examining the parameters that influenced the model significantly, the underlying pathophysiology might be better understood. Records of 1035 men containing variables including testis volume, sperm density, motility as well as the presence of endocrinopathy were randomized into 'training' and 'test' data sets. We modeled the data set using linear and quadratic discriminant function analysis, logistic regression (LR) and a neural network. Wilk's regression analysis was performed to determine which variables influenced the model significantly. Of the four models investigated, LR and a neural network performed the best with receiver operating characteristic areas under the curve of 0.93 and 0.95, respectively, correlating to a sensitivity of 28% and a specificity of 99% for the LR model, and a sensitivity and specificity of 56 and 97% for the neural network model. Reverse regression yielded P-values for the testis volume and sperm density of <0.0001. The neural network and LR models accurately predicted the probability of an endocrinopathy from testis volume, sperm density and motility without serum assays. These models may be accessed via the Internet, allowing urologists to select patients for endocrinologic evaluation at http://www.urocomp.org.

The future of sexual medicine for the urologist.

Editor's note: Winston Churchill said, 'I never worry about action, but only inaction'. Experience has taught the medical profession that action, change and adaptation are the rule as novel technologies and therapies are introduced into the mainstream of medical care. Sexual medicine is no exception. Originally thought to be psychogenic in origin, it is now well accepted that erectile dysfunction (ED) is predominately organic in origin in most middle-aged men. Treatment of organic ED has evolved with the introduction of novel, oral therapies, such as phosphodiesterase inhibitors. Adaptation has also led to incorporation of ED into the treatment realm of the primary-care physician. As sexual medicine becomes increasingly non-surgical, the challenge to the surgical specialists will reside in their ability to change and adapt to this ever-burgeoning medical discipline. Lawrence Ross, President-Elect of the American Urological Association, discusses action and change below. If urologists are to remain involved in sexual medicine, then his action plan must be brought to fruition.

Surgical sperm retrieval for assisted reproduction.

Intra-cytoplasmic sperm injection (ICSI) has revolutionized the treatment of male infertility by requiring only a single sperm to allow men whose infertility was previously considered to be uncorrectable to father a biological offspring. As a result, surgical sperm retrieval for assisted reproduction has developed to support this therapy. Microsurgical techniques have been applied to either identify areas of active spermatogenesis within the testis or to aspirate sperm-containing fluid or tissue. The combination of these techniques with in vitro fertilization (IVF)/ICSI has been shown to be a powerful approach to the treatment of azoospermic men. The availability of sperm cryopreservation offers an additional advantage, negating the need for synchronization of sperm retrieval and ovulation. Thus, the advanced methods for sperm retrieval discussed in this review also provide therapeutic options, as compared the traditional diagnostic testicular biopsy.

Apoptosis in the developing zebrafish embryo.

Apoptosis is a major part of the normal development of many organ systems and tissues. The zebrafish (Danio rerio) has become a useful model for studying early development, and recent advances in techniques used to label apoptotic cells have made it possible to visualize apoptotic cells in this model system. We have used the in situ terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) to describe the temporal and spatial distribution of apoptotic cells during normal development of the zebrafish embryo from 12 to 96 h postfertilization. By counting labeled apoptotic cells, we have demonstrated transient high rates of cell death in various structures during development, and we have correlated these peaks with previously described developmental changes in these structures. Our analysis has focused on the nervous system and associated sensory organs including the olfactory organ, retina, lens, cornea, otic vesicle, lateral line organs, and Rohon-Beard neurons. Apoptosis is also described in other non-neural structures such as the notochord, somites, muscle, tailbud, and fins.

The transporter-like protein inebriated mediates hyperosmotic stimuli through intracellular signaling.

We cloned the inebriated homologue MasIne from Manduca sexta and expressed it in Xenopus laevis oocytes. MasIne is homologous to neurotransmitter transporters but no transport was observed with a number of putative substrates. Oocytes expressing MasIne respond to hyperosmotic stimulation by releasing intracellular Ca(2+), as revealed by activation of the endogenous Ca(2+)-activated Cl(-) current. This Ca(2+) release requires the N-terminal 108 amino acid residues of MasIne and occurs via the inositol trisphosphate pathway. Fusion of the N terminus to the rat gamma-aminobutyric acid transporter (rGAT1) also renders rGAT1 responsive to hyperosmotic stimulation. Immunohistochemical analyses show that MasIne and Drosophila Ine have similar tissue distribution patterns, suggesting functional identity. Inebriated is expressed in tissues and cells actively involved in K(+) transport, which suggests that it may have a role in ion transport, particularly of K(+). We propose that stimulation of MasIne releases intracellular Ca(2+) in native tissues, activating Ca(2+)-dependent K(+) channels, and leading to K(+) transport.

Cloning of an aquaporin-like cDNA and in situ hybridization in adults of the mosquito Aedes aegypti (Diptera: Culicidae).

A cDNA encoding a putative water channel protein, aquaporin, was cloned from a cDNA library of Aedes aegypti Malpighian tubules. The cDNA encodes a 26.11 kDa protein similar to insect aquaporins from Haematobia irritans exigua (Diptera) and Cicadella viridis (Homoptera), and to mammalian aquaporin 4. Localization of the messenger RNA (mRNA) was performed by in situ hybridization of Malpighian tubules and analysed by fluorescence and confocal microscopy. The mRNA was localized in tracheolar cells associated with the Malpighian tubules. No signal was detected in the Malpighian tubule epithelium. The molecular mechanisms for water movement between tissues and tracheoles are not yet elucidated in insects. Our results suggest a model to explain fluid movements in tracheoles during insect respiration.

Steroid induction of a peptide hormone gene leads to orchestration of a defined behavioral sequence.

At the end of each molt, insects shed the old cuticle by performing preecdysis and ecdysis behaviors. Regulation of these centrally patterned movements involves peptide signaling between endocrine Inka cells and the CNS. In Inka cells, we have identified the cDNA and gene encoding preecdysis-triggering hormone (PETH) and ecdysis-triggering hormone (ETH), which activate these behaviors. Prior to behavioral onset, rising ecdysteroid levels induce expression of the ecdysone receptor (EcR) and ETH gene in Inka cells and evoke CNS sensitivity to PETH and ETH. Subsequent ecdysteroid decline is required for peptide release, which initiates three motor patterns in specific order: PETH triggers preecdysis I, while ETH activates preecdysis II and ecdysis. The Inka cell provides a model for linking steroid regulation of peptide hormone expression and release with activation of a defined behavioral sequence.

Cytosolic phospholipase A2 (cPLA2) distribution in murine brain and functional studies indicate that cPLA2 does not participate in muscarinic receptor-mediated signaling in neurons.

Cytosolic phospholipase A2 (cPLA2) catalyzes the selective release of arachidonic acid from the sn-2 position of membrane phospholipids and has been suggested as an effector in the receptor-mediated release of arachidonic acid in signal transduction. The potential role of cPLA2 as an effector in muscarinic acetylcholine receptor signaling was investigated through ectopic expression of either the m1 or m5 receptor in combination with cPLA2 in COS-1, CHO and U-373 MG cell lines. U-373 MG and COS-1 cells express undetectable or very low levels of cPLA2. CHO cell extracts are characterized by a significant endogenous PLA2 activity that was increased over 20-fold following transient expression with cPLA2 cDNA. However, in none of the cells lines did the co-expression of muscarinic receptor and cPLA2 result in a significant increase in muscarinic receptor-mediated arachidonic acid release over cells expressing muscarinic receptor alone. The distribution of cPLA2 mRNA and cPLA2 immunoreactivity in murine brain were determined in order to investigate a potential role for cPLA2 in neurotransmission. cPLA2 mRNA was expressed in white matter, including cells contained within linear arrays characteristic of interfascicular oligodendrocytes. cPLA2 immunoreactivity in white matter was evident throughout the processes of fibrous astrocytes. cPLA2 expression in gray matter was confined to astrocytes at the pial surface of the brain. cPLA2 mRNA was detected in pia mater, both at the brain surface and inner core of the choroid plexus. cPLA2 may not be directly linked to neurotransmission since enzyme expression, mRNA, and cPLA2 immunoreactivity were undetectable in neurons of murine brain. Support or regulation of neurotransmission may be provided through the activity of cPLA2 in glial cells.

Cloning of the V-ATPase B subunit cDNA from Culex quinquefasciatus and expression of the B and C subunits in mosquitoes.

The V-ATPase B subunit cDNA isolated from the midgut and Malpighian tubules of Culex quinquefasciatus larvae has a 1476 bp ORF encoding a 492 amino acids protein with a predicted mass of 54.8 kDa. Northern blot analysis reveals the presence of 1.8 and 4.2 kb transcripts in larvae and a 3.0 kb transcript in pupae. A single 57 kDa protein band is detected in immunoblot analysis of protein extracts from C. quinquefasciatus and A. aegypti larvae. Using antibodies to the B subunits we demonstrate high-level expression of these subunits in ion-transporting cells of caeca and anterior midgut, and in Malpighian tubules and rectum. High V-ATPase expression was also observed in the larval salivary glands, central nervous system neurophile, the thoracic endocrine complex and in imaginal discs.

Isolation of the V-ATPase A and c subunit cDNAs from mosquito midgut and Malpighian tubules.

Using conserved amino acid sequences for the design of oligonucleotide primers, we isolated cDNA clones for two subunits of the V-ATPase from the midgut and Malpighian tubules of Aedes aegypti larvae. The 3.1 kb cDNA of the A subunit of the peripheral catalytic V1 sector codes for a protein of 68.6 kDa. The protein contains conserved motifs, including an ATP/GTP binding site, found in all other A subunits. Southern analysis using the A subunit as a probe suggests the presence of only a single copy of gene in the Aedes aegypti. The 0.85 kb cDNA of the c subunit of the membrane H+ conducting V0 sector codes for a protein of kDa. This protein has four transmembrane domains and contains a conserved glutamic acid that serves as the binding site for dicyclohexylcarbodiimide. Southern analysis using the c subunit as a probe suggests the presence of more than one related gene in the genome of Aedes aegypti. Pileup analysis of various A and c subunits shows that these subunits fall into distinct clusters, including one in which all arthropod proteins are clustered.

Cloning, functional expression, and pharmacology of a GABA transporter from Manduca sexta.

Termination of synaptic transmission occurs by several mechanisms that include uptake of the neurotransmitter molecules into the presynaptic neuron by specialized membrane transport proteins. We have cloned a (DABA)-sensitive gamma-aminobutyric acid (GABA) transporter from a cDNA library from Manduca sexta embryo. The cDNA clone, MasGAT, shows high sequence homology to known mammalian GABA transporters. The transcript is about 5.5 kb with an open reading frame of 1793 bp. Injection of a 2.2-kb cRNA from this clone into Xenopus oocytes results in [3H]GABA transport. A Michaelis-Menten kinetic analysis shows that GABA transport occurs by a high-affinity and saturable process, suggesting that it is carrier-mediated. Ion substitution studies also show the transport process to be highly dependent on extracellular Na+ gradient, a finding that is consistent with properties of known mammalian neurotransmitter transporters. Although MasGAT shares certain pharmacological similarities with known mammalian GABA transporters, this transporter is pharmacologically distinct from the known mammalian GABA transporters.

Varicocele vein ligation in 565 patients under local anesthesia: a long-term review of technique, results and complications in light of proposed management by laparoscopy.

Several recent reports have suggested that laparoscopic internal spermatic vein ligation provides a simpler, less debilitating and more cost efficient method of varicocele ligation than conventional surgical techniques. We analyzed the results of open varicocele ligation using local anesthesia in 565 patients for 10 years. All surgery was performed in the outpatient setting using 0.5% lidocaine. In most patients 50 to 200 mcg. fentanyl or 3 to 7 mg. midazolam were used for intravenous sedation. The average operating time, including the administration of anesthesia, was 39 minutes for unilateral and 71 minutes for bilateral procedures. All patients returned to light duty work in 24 to 48 hours and full strenuous physical activity within 1 week. The only complications encountered were 2 wound hematomas (0.3%), 4 minor wound separations (0.7%) and 41 hydroceles (7.3%). Semen improvement and pregnancy rates were similar to those reported in prior series. This study demonstrates that varicocele vein ligation can be done rapidly, efficiently and safely using local anesthesia with time of recovery and return to work comparable to those reported for laparoscopic techniques.

Microsurgical epididymovasostomy: predictors of success.

The results of microsurgical epididymovasostomy for congenital and acquired vasoepididymal obstruction were retrospectively reviewed in 22 patients in an attempt to determine what preoperative or intraoperative factors might predict surgical success. The overall success rate, defined as sperm on postoperative semen analysis, was 48%. The presence of sperm on an intraoperative touch preparation from the epididymis was significantly correlated with response (chi-square 3.24, p < 0.10) and no patient without sperm on touch preparation had sperm on subsequent semen analyses. Testicular biopsy positive for spermatogenesis and presence of motile sperm on intraoperative touch preparation were not statistically significant predictors of response. These results suggest that presence or absence of sperm on intraoperative touch preparation is the only significant prognosticator of successful microsurgical epididymovasostomy.

Initial tract formation in the mouse brain.

Mouse embryos from embryonic days 8.5-10.5 (E8.5-E10.5) were fixed and labeled with an antibody to neuron-specific class III beta-tubulin (Moody et al., 1987; Lee et al., 1990a,b) to reveal the first neurons, axons, and tracts in the brain. They were studied in whole-mounts and in light microscopic sections. Some conclusions were checked by labeling tracts in older embryos (E11.5 and E12.5) with the lipophilic dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine. The first immunoreactive cells appeared at E8.5, prior to neural tube closure, in the neural plate immediately caudal to the optic vesicle. Cells along the dorsal midline of the mesencephalon issued the first axons, on E9.0; the cells were the mesencephalic nucleus of the trigeminal nerve, and the axons formed its descending tract. The tract reached the level of the trigeminal ganglion by E10.0 but did not enter the ganglion until after E12.5. On E9.5, the number of labeled cells and axons in the alar plate of the presumptive diencephalon and mesencephalon had increased substantially, and many of the rostral ones coursed into the basal plate to enter longitudinal tracts there. Two tracts originated from cells in the basal plate: the tract of the postoptic commissure (from the base of the optic stalk to the level of the cephalic flexure) and the medial longitudinal fasciculus (from the level of the cephalic flexure caudally through the mid and hind-brains). By E10.0, a small mammillotegmental tract paralleled the tract of the postoptic commissure, but immunolabeling was so widespread that discrete tracts were impossible to discern in the presumptive diencephalon and mesencephalon. The more rostral regions remained lightly labeled. In the cerebral vesicle, the presumptive cerebral cortex, the first immunoreactive cells appeared at E10.0; they had multiple processes oriented parallel to the pia, and were identified as the Cajal-Retzius cells. By E10.5, no tracts had formed in the cerebral vesicle. All the tracts formed by E10.0 were superficial, in the subpial lamina. Those that can be identified in the adult brain are very deep structures. These results are compared with previous descriptions of the embryonic brains of amphibians, fish, birds, and other mammals, including humans.