Micro-fabricated perforated polymer devices for long-term drug delivery

Fabrication techniques have been developed to produce a perforated polymer microtube as a drug delivery device. The technique consists of first forming a silicon platform with trenches and alignment marks to hold the tubes for subsequent processing. Photolithography and reactive ion etching with an inductively coupled plasma source were used to fabricate micro holes on the surface of polyimide tubes. Several materials have been used to form the etching mask, including titanium film deposited by e-beam evaporation and SiO₂ and SiN_x films deposited by high-density plasma chemical vapor deposition (HDPCVD). Three equidistant holes of 20 μm in diameter were fabricated on polyimide tubes (I.D. = 125 μm). The perforated tubes were loaded with ethinyl estradiol and tested for drug release in phosphate buffered saline (pH = 7.1) at 37°C. Zero order release was observed over a period of 30 days with a potential to be extended to 4 years.     

Microsporidian parasites: a danger facing marine fishes of the Red Sea

Out of 600 marine fish from the Red Sea belonging to three different species that were collected and examined for microsporidian parasites, 87 (14.5%) fish were found to be infected. The infection was recorded as cysts or xenomas embedded in the gut epithelium and the peritoneal cavity of the three fish species. The highest percent of infection with microsporidian parasites was recorded in Saurida tumbil 19.5% (39/200) followed by Pagrus pagrus 15% (45/300) and the lowest percent of infection was recorded in Epinephelus chlorostigma 3% (three out of 100). After rupture of the cysts, the spores were released and examined by light microscopy. Each spore was elongated to ellipsoidal in shape and possessed a posterior vacuole which is characteristic to phylum Microspora. They measure 1.6 ± 0.5 μm (1.5–2.4 μm) × 1.3 ± 0.1 μm (1.3–2.0 μm) in Saurida tumbil and Pagrus pagrus, respectively. The spores of Pleistophora sp recorded from E. chlorostigma were ovoid to pyriform in shape and measure 1.9 ± 0.5 μm (1.8–2.7 μm) × 1.6 ± 0.4 μm (1.5–2.4 μm).     

RNAi-mediated silencing of a novel Ascaris suum gene expression in infective larvae

In the present study, the potential of RNA interference (RNAi) as a gene silencing tool and the resultant effects on Ascaris suum larval development was examined by targeting a gene (represented by the EST 06G09) specifically expressed in the infective larvae of A. suum. BALB/c mice were infected with RNAi-treated larvae. The results showed that the target gene was silenced after soaking for 72 h, and the survival rate of the RNAi-treated larvae was reduced by 17.25% (P < 0.01). A significant difference (P < 0.05) was detected in the numbers of larvae collected from the livers and lungs of infected mice 4 days after infection with untreated larvae (164.29 ± 21.51) and RNAi-treated larvae (71.43 ± 14.35). Significant differences (P < 0.01) were also found in the body length and width between untreated larvae (480 ± 105.77 μm for length and 23.93 ± 3.72 μm for width) and RNAi-treated larvae (400.57 ± 71.31 μm for length and 20.20 ± 2.43 μm for width). These results show that the gene represented by EST 06G09 may play a role in the development of A. suum larvae.     

<Emphasis Type="Italic">Myxobolus honghuensis</Emphasis> n. sp. (Myxosporea: Bivalvulida) parasitizing the pharynx of allogynogenetic gibel carp <Emphasis Type="Italic">Carassius auratus gibelio</Emphasis> (Bloch) from Honghu Lake,China

Myxobolus honghuensis n. sp. is described from allogynogenetic gibel carp Carassius auratus gibelio (Bloch), during a survey of myxosporean parasites in Honghu Lake, Hubei Province, China. It is characterized by the presence of round plasmodia of 5–12 mm in diameter in the pharynx of host. Mature spores of M. honghuensis were pyriform in frontal view and anterior pointed with bluntly round posterior, they measured 16.9 ± 0.5 (15.1–19.5) μm long, 10.4 ± 0.4 (9.0–11.3) μm wide, and 8.4 ± 0.4 (7.9–9.1) μm thick. Two polar capsules were pyriform and slightly unequal with larger polar capsule 8.4 ± 0.4 (7.6–10.2) μm × 3.9 ± 0.2 (3.0–4.5) μm and smaller capsule 7.9 ± 0.2 (7.0–9.3) μm × 3.7 ± 0.3 (2.8–4.1) μm. Polar filaments coiled with seven to eight turns. Both morphology and DNA sequence data revealed that M. honghuensis n. sp. was distinct from other described Myxobolus species. Phylogenetic analysis placed M. honghuensis n. sp. in a clade of gill-infecting myxobolids.     

Insight in Human Skin Microcirculation Using In Vivo Reflectance-Mode Confocal Laser Scanning Microscopy

Reflectance-mode confocal laser scanning microscopy allows in vivo imaging of the human skin. We hypothesized that this high-resolution technique enables observation of dynamic changes of the cutaneous microcirculation. Twenty-two volunteers were randomly divided in two groups. Group 1 was exposed to local heating and group 2 to local cold stress. Confocal microscopy was performed prior t ₀ (control), directly t ₁ and 5 min t ₂ after local temperature changes to evaluate quantitative blood cell flow, capillary loop diameter, and density of dermal capillaries. In group 1, blood flow increased at t ₁ (75.82 ± 2.86/min) and further at t ₂ (84.09 ± 3.39/min) compared to the control (61.09 ± 3.21/min). The control capillary size was 9.59 ± 0.25 μm, increased to 11.16 ± 0.21 μm (t ₁) and 11.57 ± 0.24 μm (t ₂). The dermal capillary density increased in t ₁ (7.26 ± 0.76/mm²) and t ₂ (8.16 ± 0.52/mm²), compared to the control (7.04 ± 0.62/mm²). In group 2, blood flow decreased at t ₁ (41.73 ± 2.61/min) and increased at t ₂ (83.27 ± 3.29/min) compared to the control (60.73 ± 2.90/min). The control capillary size was 9.55 ± 0.25 μm, decreased at t ₁ (7.78 ± 0.26 μm) and increased at t ₂ (11.38 ± 0.26 μm). Capillary density decreased at t ₁ (5.01 ± 0.49/mm²) and increased at t ₂ (7.28 ± 0.53/mm²) compared to the control (7.01 ± 0.52/mm²). Confocal microscopy is a sensitive and noninvasive imaging tool for characterizing and quantifying dynamic changes of cutaneous microcirculation on a histomorphological level.     

Afferent arteriolopathy and glomerular collapse but not segmental sclerosis induce tubular atrophy in old spontaneously hypertensive rats

In chronic renal disease, the temporal and spatial relationship between vascular, glomerular and tubular changes is still unclear. Hypertension, an important cause of chronic renal failure, leads to afferent arteriolopathy, segmental glomerulosclerosis and tubular atrophy in the juxtamedullary cortex. We investigated the pathological changes of hypertensive renal disease in aged spontaneously hypertensive rats using a large number of serial sections, where we traced and analyzed afferent arteriole, glomerulus and proximal tubule of single nephrons. Our major finding was that both afferent arteriolopathy and glomerular capillary collapse were linked to tubular atrophy. Only nephrons with glomerular collapse (n = 13) showed tubules with reduced diameter indicating atrophy [21.66 ± 2.56 μm vs. tubules in normotensive Wistar Kyoto rats (WKY) 38.56 ± 0.56 μm, p < 0.05], as well as afferent arteriolar wall hypertrophy (diameter 32.74 ± 4.72 μm vs. afferent arterioles in WKY 19.24 ± 0.98 μm, p < 0.05). Nephrons with segmental sclerosis (n = 10) did not show tubular atrophy and tubular diameters were unchanged (35.60 ± 1.43 μm). Afferent arteriolar diameter negatively correlated with glomerular capillary volume fraction (r = −0.36) and proximal tubular diameter (r = −0.46) implying reduced glomerular and tubular flow. In line with this, chronically damaged tubules showed reduced staining for the ciliary protein inversin indicating changed ciliary signalling due to reduced urinary flow. This is the first morphological study on hypertensive renal disease making correlations between vascular, glomerular and tubular components of individual nephron units. Our data suggest that afferent arteriolopathy leads to glomerular collapse and reduced urinary flow with subsequent tubular atrophy.     

Effects of K-201 on the calcium pump and calcium release channel of rat skeletal muscle

The benzothiazepine derivative K-201 has been suggested as a potential therapeutic agent due to its antiarrhythmogenic action. To understand how the drug alters calcium release from the sarcoplasmic reticulum (SR), we investigated its effects on the SR calcium channel and calcium pump by single channel electrophysiology, whole-cell confocal microscopy, and ATPase activity measurements on control and post-myocardial infarcted (PMI) rat skeletal muscle. In bilayers, K-201 induced two subconductance states corresponding to ∼24% (S₁) and ∼13% (S₂) of the maximum conductance. Dependence of event frequency and of time spent in S₁ and S₂ on the drug concentration was biphasic both in control and in PMI rats, with a maximum at 50 μM. At this concentration, the channel spends 26 ± 4% and 24 ± 4%, respectively, of the total time in these subconductance states at positive potentials, while no subconductances are observed at negative potentials. K-201 altered the frequency of elementary calcium release events: spark frequency decreased from 0.039 ± 0.001 to 0.023 ± 0.001 s⁻¹ sarcomere⁻¹, while the frequency of embers increased from 0.011 ± 0.001 to 0.023 ± 0.001 s⁻¹ sarcomere⁻¹. Embers with different amplitude levels were observed after the addition of the drug. K-201 inhibited the Ca²⁺ ATPase characterized by IC_50,contr = 119 ± 21 μM and n _Hill,contr = 1.84 ± 0.48 for control and IC_50,PMI = 122 ± 18 μM and n _Hill,PMI = 1.97 ± 0.24 for PMI animals. These results suggest that although K-201 would increase the appearance of subconductance states, the overall calcium release is reduced by the drug. In addition, the effect of K-201 is identical on calcium release channels from control and PMI rats.     

Effect of praziquantel prolonged administration on granuloma formation around Schistosoma japonicum eggs in lung of sensitized mice

Schistosomiasis remains a major public health problem and it is an immune disease. The schistosome egg is the primary parasite factor responsible for the overt disease. The eggs release the soluble antigen, which induces intensive tissue reaction, a granulomatous reaction to the eggs. If granuloma formation could be suppressed, overt disease might not develop. Praziquantel is an effective antischistosomal drug especially for adult worms. However, whether praziquantel has a suppressing effect on granuloma formation around schistosome eggs directly remains unclear. The purpose of the present study was to investigate the effect of praziquantel, especially administered persistently, on granuloma formation around Schistosoma japonicum eggs in the lung of sensitized mice. Thirty-six mice were divided into three groups averagely. Group A was a control group. First, the mice were injected with schistosomal eggs hypodermically in abdomen, and 10 days later injected with schistosomal eggs intravenously via a tail vein. Group B was a praziquantel short administration group. In addition to the injections of schistosomal eggs as the same of Group A, the mice were administered with praziquantel in a daily dose of 300 mg/kg for 3 days, from 1 day before the intravenous injection of the eggs. Group C was a praziquantel prolonged administration group. In addition to the injections of schistosomal eggs as the same of Group A, the mice were administered with praziquantel in a daily dose of 150 mg/kg for 5 days weekly until the mice were sacrificed. Three mice of each group were sacrificed on days 7, 14, 28, and 56, respectively after the intravenous injection of the eggs, and the lung tissues were fixed with formalin and the slices were HE stained. The granulomas containing eggs in their centers were selected, and 25–30 granulomas from the animals of each group were measured at each time period. The mean areas of egg granulomas of each group were calculated, and the neutrophilic granulocytes, eosinocytes, lymphocytes, fibroblasts, and macrophages within the egg granulomas were counted. The mean numbers of them of each group were calculated. All the data of each group were analyzed and compared statistically. On day 56 after the intravenous injection of the eggs, the mean area of schistosomal egg granulomas in group B was (227.4 ± 728.0) × 10³ μm², less than that of [(297.9 ± 153.3) × 10³ μm²] in group A, and the suppression rate was 23.7% (P < 0.05). On days 7, 14, 28, and 56, the mean areas of schistosomal egg granulomas in group C were (575.8 ± 155.6) × 10³ μm², (310.5 ± 854.0) × 10³ μm², (267.7 ± 513.3) × 10³ μm², and (214.9 ± 446.4) × 10³ μm², respectively, significantly less than those of [(692.7 ± 232.6) × 10³ μm², (439.4 ± 165.0) × 10³ μm², (385.7 ± 129.3) × 10³ μm², and (297.9 ± 153.3) × 10³ μm²] in group A. The suppression rates were 16.9%, 29.3%, 30.6%, and 27.9%, respectively (P values <0.05). On day 56, the mean numbers of neutrophilic granulocytes were 11.4 ± 5.0 in group A and 5.2 ± 3.1 in group C, respectively, with the suppression rate of 54.4% in group C (P < 0.05). On day 56, the mean numbers of eosinocytes within the egg granulomas were 2.3 ± 2.0, 0.1 ± 0.3, and 0.3 ± 0.6 in groups A, B, and C, respectively, with the suppression rate of 95.7% in group B and 87.0% in group C (P values <0.05). On day 56, the mean numbers of macrophages within egg granulomas were 14.3 ± 6.9 in group C, compared with 18.6 ± 8.2 in group A, the suppression rate was 23.1% (P < 0.05). On day 56, the mean numbers of fibroblasts within the egg granulomas were 6.6 ± 4.4 and 5.8 ± 2.6 in groups B and C, respectively, and compared with 14.3 ± 7.8 in group A, the increasing extents decreased by 53.8% and 59.4%, respectively (P values <0.05). Therefore, the administration of praziquantel, especially the prolonged administration, can suppress the formation of schistosomal egg granulomas, including reduction in the areas of granulomas and suppression of the inflammatory cells and the hyperplasia of fibroblasts within granulomas.     

The effects of interleukin (IL)-12 and IL-4 deficiency on worm development and granuloma formation in Schistosoma japonicum-infected mice

CD4⁺ T-helper (Th) cell is widely recognized to be capable of influencing worm development and egg granuloma formation after schistosome infection. Interleukin (IL)-12 and IL-4 play key roles in regulation of Th cell differentiation. In the present study, we subcutaneously inoculated mice with hybridoma cells secreting monoclonal antibodies to neutralize IL-12 and IL-4 and explored the effects of IL-12 and IL-4 deficiency on the worm development and granuloma formation in mice infected with cercariae of Schistosoma japonicum. It was found that deficiency of host IL-12 and IL-4 supported normal parasite survival and fecundity. However, worm development (length and female fecundity) was significantly enhanced in anti-IL-12-treated mice. Mean length of worms in anti-IL-12-treated group was significantly greater than that of intact controls on day 28 after infection (females, 11.84 ± 1.20 mm vs. 9.45 ± 1.34; males, 9.35 ± 1.21 mm vs. 8.10 ± 0.85 mm, p < 0.05). Liver egg load per pair of worms (1,770.12 ± 470.67 vs. 806.08 ± 232.37, p < 0.05) and uterine egg load of ovigerous females (93.08 ± 27.85 vs. 46.05 ± 34.24, p < 0.05) in anti-IL-12-treated mice were significantly higher than those in intact control 28 days postinfection. But these effects diminished 42 days postinfection (p > 0.05). Granuloma size in anti-IL-12-treated mice was significantly larger than that in intact mice 42 days postinfection (398.3 ± 80.7 μm vs. 294.4 ± 72.2 μm, p < 0.05). Granuloma fibrosis dramatically intensified in anti-IL-12-treated mice but diminished in anti-IL-4-treated mice. The results suggest that IL-12 may play an impeditive role in the development of S. japonicum and in granuloma formation as well as fibrosis. IL-4 may promote granuloma formation but have no effect on worm development.     

Exenatide enhances kaliuresis under conditions of hyperkalemia

Experiments on Wistar rats showed that exenatide (0.015–0.5 nmol per 100 g body weight) somewhat increased renal excretion of potassium from 7 ± 1 to 16 ± 1 μmol/h/100 g body weight (p < 0.05) in animals with normal serum concentration of glucose (4.6 ± 0.4 mM) and potassium (4.3 ± 0.1 mM). Exenatide dramatically enhanced excretion of potassium under conditions of hyperkalemia (11.4 ± 0.4 mM) produced by intraperitoneal injection of 1.25% KCl solution (5 ml per 100 g body weight). During the fi rst postinjection hour, potassium excretion increased 2-fold and attained 97 ± 11 μmol/h/100 g body weight in comparison with potassium load alone (47 ± 9 μmol/h/100 g body weight, p < 0.05). The data attest to a possible role of peptide regulators in normalization of potassium balance via renal mechanisms.     

A novel herbal formulation against dengue vector mosquitoes <Emphasis Type="Italic">Aedes aegypti</Emphasis> and <Emphasis Type="Italic">Aedes albopictus</Emphasis>

The objective of this study was to develop a herbal formulation to control dengue vector mosquitoes. PONNEEM, a novel herbal formulation prepared using the oils of neem (Azadirachta indica), karanj (Pongamia glabra) and their extracts, was tested for larvicidal, ovicidal and oviposition deterrent activities against Aedes aegypti and Aedes albopictus at 1, 0.5, 0.3 and 0.1 ppm concentrations. Cent percent larvicidal and ovicidal activities were observed at 0.1 ppm in the two mosquito species under laboratory and sunlight-exposed conditions up to 12 months from the date of manufacture. Oviposition deterrent activity of 69.97% and 71.05% was observed at 1 ppm concentration of PONNEEM against A. aegypti and A. albopictus, respectively. Reduction in enzyme levels for α-esterase was 0.089 ± 0.008 and 0.099 ± 0.140 μg napthol produced/min/mg larval protein; for β-esterase, it was 0.004 ± 0.009 and 0.001 ± 0.028 μg napthol produced/min/mg larval protein; for glutathione S-transferase, it was 10.4814 ± 0.23 and 11.4811 ± 0.21 μmol/min/mg larval protein and for total protein, it was 0.177 ± 0.010 and 0.008 ± 0.005 mg/individual larva in treated groups of A. aegypti and A. albopictus, respectively. The nontarget organisms such as Gambusia affinis and Diplonychus indicus were not affected. No mortality was observed in control. PONNEEM can be used effectively for the management of human vector mosquitoes.     

Temperature Pretreatment Alters the Polarization Response of Human Neutrophils to the Chemoattractant N-formyl-Met-Leu-Phe

Neutrophils present a polarized morphology upon stimulation of chemoattractants, which play a vital role in host-defense mechanisms. Many studies have been published on neutrophil polarization, in which three different temperatures pretreatment (4°C, 25°C and 37°C) have been used. However, no study has investigated whether different temperature pretreatments affect neutrophil polarization. In the current study, we examined the effects of 4°C, 25°C and 37°C pretreatment temperatures on short-term (1 or 3 min) chemoattractant-induced polarization. Human neutrophils were polarized upon the stimulation of N-formyl-Met-Leu-Phe (fMLP) after pretreated by different temperature. The morphological changes of the neutrophils were investigated under the microscopy. The F-actin polymerization was determined by immunological histological chemistry. There were more head–tail polarized cells (>50% of the cells) in the 25°C and 37°C pretreatment groups than in the 4°C group (32.4%). The average lengths of the pseudopod were 3.2 ± 1.1 μm (n = 17), 5.3 ± 2.1 μm (n = 12) and 7.4 ± 2.7 μm (n = 21) in the 4°C, 25°C and 37°C pretreatment groups, respectively; the 4°C and 37°C pretreatment groups were statistically different (P < 0.05). Additionally, there was a statistically significant difference in the pseudopod extension rate among the three groups, as well as the Lamellipod percentage between the 4°C group and the other two groups within 1 min of stimulation with fMLP. This study demonstrates that different temperature pretreatments affect neutrophil polarization upon short-term stimulation with fMLP. Dongliang Zhao, Xiaojing Meng, Chunqing Cai equally contributed to this paper.     

ATP Concentration in the Cingulum Bundle of Rats during Stimulation of the Ventromedial Hypothalamus

We measured the concentration of ATP in the posterior cingulum bundle of Sprague-Dawley rats during electrical stimulation of the negative emotiogenic zone in the ventromedial hypothalamus. Electrostimulation of the ventromedial hypothalamus in animals was accompanied by an increase in systolic and diastolic blood pressure, which illustrates the autonomic response to this treatment. Variations in BP of rats were followed by an increase in ATP content in the posterior cingulum bundle. After stimulation of the ventromedial hypothalamus, ATP concentration in the cingulum bundle reached the maximum levels of 1.3 ± 0.3 μM (n = 4) and 1.67 ± 0.43 μM (n = 10) and remained high for 42.6 ± 7.5 sec. The enhanced release of ATP in the cingulate region of the brain is probably related to the involvement of this structure into emotional reactions. ATP plays a role of the major energy source and signal molecule, which provides the adequate biochemical and physiological processes and cell-to-cell interaction in CNS upon the exposure to negative emotiogenic factors.     

Transmural Oxygen Tension Gradients in Rat Cerebral Cortex Arterioles

Modified needle oxygen microelectrodes and vital microscopy were used to measure transmural oxygen tension gradients (PO₂) in pial arterioles with lumen diameters of 20–90  μm. A relationship between the magnitude of the transmural PO₂ gradient and arteriole wall tone was found: in control conditions, PO₂ gradients were 1.17 ± 0.06 mmHg/μm (n  = 40), while in conditions of arteriolar wall dilation the transmural PO₂ gradient decreased to 0.68 ± 0.04 mmHg/μm (p  <  0.001, n  = 38). These data provide the first measurements of transmural PO₂ gradients in pial arterioles of different calibers at different levels of vascular tone and have fundamental importance for assessing the role of arterial microvessels in tissue oxygen supply processes. The results obtained here provide evidence that oxygen consumption by the vessel wall is within the range characteristic of enveloping tissues and that oxygen consumption by the endothelial cell layer probably has no significant effect on the magnitude of the transmural PO₂ gradient. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 94, No. 4, pp. 394–405, April, 2008.     

Haematology and serum biochemistry of golden eagle (Aquila chrysaetos) in Iran

Haematological and serum biochemical values were estimated in blood samples collected from 21 apparently adult golden eagles (Aquila chrysaetos) of both sexes. The mean values of red blood cells, packed cell volume, haemoglobin, white blood cells, heterophils, lymphocytes, monocytes and eosinophils were 1.63 ± 0.11 × 10¹²/l, 0.47 ± 0.009 l/l, 91.73 ± 1.52 g/l, 24.31 ± 1.97 × 10⁹/l, 4.40 ± 0.22 × 10⁹/l, 16.81 ± 0.65 × 10⁹/l, 0.99 ± 0.19 × 10⁹/l and 2.10 ± 0.30 × 10⁹/l, respectively. The leucocytes had 69.14%, 4.09%, 18.12% and 8.65% lymphocytes, monocytes, heterophils and eosinophils, respectively. The results of serum biochemistry in the golden eagle indicated that the concentrations of glucose, total protein, albumin, total globulin, cholesterol, triglyceride, uric acid, blood urea nitrogen, creatinine, calcium, phosphorous, aspartate aminotransferase, alanine aminotransferase, creatine kinase, lactate dehydrogenase and alkaline phosphatase were 16.42 ± 0.73 mmol/l, 49.76 ± 1.35 g/l, 20.46 ± 0.79 g/l, 29.30 ± 1.47 g/l, 2.14 ± 0.09 mmol/l, 2.04 ± 0.08 mmol/l, 457.67 ± 97.46 μmol/l, 2.74 ± 0.17 mmol/l, 53.27 ± 3.87 μmol/l, 2.37 ± 0.24 mmol/l, 1.73 ± 0.08 mmol/l, 293.24 ± 18.96 IU/l, 28.21 ± 2.36 IU/l, 411.29 ± 58.37 IU/l, 1,209.89 ± 21.73 IU/l and 67.31 ± 5.29 IU/l, respectively. There were no significant differences between haematological and serum biochemical parameters of male and female golden eagles (P > 0.05).     

Trisk 32 regulates IP(3) receptors in rat skeletal myoblasts

To date, four isoforms of triadins have been identified in rat skeletal muscle. While the function of the 95-kDa isoform in excitation–contraction coupling has been studied in detail, the role of the 32-kDa isoform (Trisk 32) remains elusive. Here, Trisk 32 overexpression was carried out by stable transfection in L6.G8 myoblasts. Co-localization of Trisk 32 and IP₃ receptors (IP₃R) was demonstrated by immunocytochemistry, and their association was shown by co-immunoprecipitation. Functional effects of Trisk 32 on IP₃-mediated Ca²⁺ release were assessed by measuring changes in [Ca²⁺]_i following the stimulation by bradykinin or vasopressin. The amplitude of the Ca²⁺ transients evoked by 20 μM bradykinin was significantly higher in Trisk 32-overexpressing (p < 0.01; 426 ± 84 nM, n = 27) as compared to control cells (76 ± 12 nM, n = 23). The difference remained significant (p < 0.02; 217 ± 41 nM, n = 21, and 97 ± 29 nM, n = 31, respectively) in the absence of extracellular Ca²⁺. Similar observations were made when 0.1 μM vasopressin was used to initiate Ca²⁺ release. Possible involvement of the ryanodine receptors (RyR) in these processes was excluded, after functional and biochemical experiments. Furthermore, Trisk 32 overexpression had no effect on store-operated Ca²⁺ entry, despite a decrease in the expression of STIM1. These results suggest that neither the increased activity of RyR, nor the amplification of SOCE, is responsible for the differences observed in bradykinin- or vasopressin-evoked Ca²⁺ transients; rather, they were due to the enhanced activity of IP₃R. Thus, Trisk 32 not only co-localizes with, but directly contributes to, the regulation of Ca²⁺ release via IP₃R.     

Phylogenetic characterization of a microsporidium (Endoreticulatus sp. Zhenjiang) isolated from the silkworm, Bombyx mori

This study examined the morphological and molecular characteristics of the microsporidium Endoreticulatus sp. Zhenjiang, isolated from the silkworm (Bombyx mori). The fresh spores were oval, 2.9 ± 0.2 μm in length and 1.2 ± 0.2 μm in width. The complete rRNA cistron has a length of 4,432 bp (GenBank accession no. FJ772431), including the large subunit rRNA (2,460 bp), the internal transcribed spacer (187 bp), the small subunit rRNA (1,254 bp), the intergenic spacer (276 bp), and the 5S region (115 bp). The organization of the rRNA gene is 5′-LSU-ITS-SSU-IGS-5S-3′, which is reverse compared to the organization of most microsporidian rRNA regions. Phylogenetic analysis based on small subunit rRNA sequences showed that this isolate belongs to the genus Endoreticulatus, and is closely related to Glugoides intestinalis. Furthermore, both had a similar reverse arrangement of the rRNA gene. Our study provides another example of a microsporidian species with a novel organization of rRNA genes, demonstrating that the reverse arrangement is exhibited not only by the microsporidian genus Nosema but may also occur in a clade that contains the genera Endoreticulatus and Glugoides.     

Two new species of Myxobolus (Myxozoa: Myxosporea: Bivalvulida) infecting Indian freshwater fishes in Punjab Wetlands (India)

A survey of parasites of freshwater fishes in Harike and Ropar Wetlands of Punjab (India) revealed the presence of two new myxosporean species belonging to the genus Myxobolus Butschli 1882. Spores of the first species, M. duodenalis sp. nov. parasitize the inner wall of the duodenum of Wallago attu (Bloch and Schneider) (Cypriniformis: Cyprinidae) vern. mulli are characterized by a broad, pyriform shape with a blunt anterior end and measures 9.0 × 3.20 μm. Spore valves are thin, smooth, and symmetrical with no parietal folds. There are two polar capsules, prominently unequal and pyriform in shape, with a narrow anterior end and a rounded posterior end. The larger polar capsule measures 4.70 × 1.76 μm and the smaller 2.76 × 1.06 μm. The larger polar capsule occupies more than half, while the smaller one occupies one third of the spore body cavity. An intercapsular process is absent. Spores of the second species, Myxobolus patialensis sp. nov. parasitize the caudal fin of Labeo rohita (Ham. 1822) (Cypriniformis: Cyprinidae) vern. rohu are pyriform in valvular view, measuring 11.28 × 6.67 μm. The two shell valves are asymmetrical and contain five parietal folds along the posterior end of the spore. Two anteriorly situated polar capsules, elongated and oval in shape are prominently unequal in size. The larger polar capsule measures 4.8 × 3.1 μm and smaller one 1.70 × 1.51 μm. The larger polar capsule lies obliquely to the spore axis and the smaller one is at the same level but pointing outward anteriolaterally. An intercapsular process is absent. This species is characterized by having a prominent ridge on the shell surface anteriolaterally on the side of the smaller polar capsule.     

Design considerations in the development and application of microdisc electrode arrays (MDEAs) for implantable biosensors

The use of microlithographically fabricated Microdisc Electrode Arrays (MDEAs) in the development of implantable voltammetric biosensors necessitates design criteria that balances the overall footprint of the device with the advantages to be derived from large separation distances between non-interacting microdisc elements. Using the dynamic electroanalytical techniques of Multiple Scan Rate Cyclic Voltammetry (MSRCV) experiments with finite element simulations and Electrochemical Impedance Spectroscopy with equivalent circuit modeling, three unique MDEA designs; MDEA 050 (r = 25 μm, 5,184 discs), MDEA 100 (r = 50 μm, 1,296 discs) and MDEA 250 (r = 125 μm, 207 discs) of constant critical dimensions (center-to-center d/r = 4) and area (A = 0.1 cm²) were studied in 1.0 mM ferrocene monocarboxylic acid (FcCO₂H) solution (in 0.1 M Tris/0.1 M KCl buffer, pH = 7.2). The critical disc-to-disc spacing (d/r) required to archive 67% of maximal current response was defined as optimal. Based on the predictive model, new MDEA designs; MDEA 001 (r = 0.5 μm, 127,324 discs), MDEA 002.5 (r = 1.25 μm, 20,372 discs), MDEA 005 (r = 2.5 μm, 5,093 discs), MDEA 010 (r = 5 μm, 1,273 discs), MDEA 015 (r = 7.5 μm, 566 discs), MDEA 020 (r = 10 μm, 318 discs) were simulated at 10 and 100 mV/s. The final disc count of each MDEA was dictated by the need to maintain a comparable electroactive area between the MDEAs, which was chosen to be 0.001 cm², which in turn was dictated by the need to generate sufficient electrochemical current to be comfortably measured by common electrochemical detectors.