A randomised controlled feasibility trial of a clinical protocol to manage hypotension during major non-cardiac surgery.

Intra-operative hypotension is a risk factor for postoperative morbidity and mortality. Minimally invasive monitors that derive other haemodynamic parameters, such as stroke volume, may better inform the management of hypotension. As a prelude to a multicentre randomised controlled trial, we conducted a single-centre feasibility trial of a protocol to treat hypotension as informed by minimally invasive haemodynamic monitoring during non-cardiac surgery. We recruited adults aged ≥40 years with cardiovascular risk factors who underwent non-cardiac surgery requiring invasive arterial pressure monitoring. Participants were randomly allocated to usual care, or a clinical protocol informed by an arterial waveform contour analysis monitor. Participants, outcome assessors, clinicians outside operating theatres and analysts were blinded to treatment allocation. Feasibility was evaluated based on: consent rate; recruitment rate; structured feedback from anaesthesia providers; and between-group differences in blood pressure, processes-of-care and outcomes. The consent rate among eligible patients was 33%, with 30 participants randomly allocated to the protocol and 30 to usual care. Anaesthesia providers rated the protocol to be feasible and acceptable. The protocol was associated with reduced fluid balance and hypotension exposure in the peri-operative setting. Postoperative complications included: acute myocardial injury in 18 (30%); acute kidney injury in 17 (28%); and surgical site infection in 7 (12%). The severity of complications was rated as moderate or severe in 25 (42%) participants. In summary, this single-centre study confirmed the feasibility of a multicentre trial to assess the efficacy and safety of a physiologically guided treatment protocol for intra-operative hypotension based on minimally invasive haemodynamic monitors.

High volatile anaesthetic conservation with a digital in-line vaporizer and a reflector.

BACKGROUND: A volatile anaesthetic (VA) reflector can reduce VA consumption (VAC) at the cost of fine control of its delivery and CO2 accumulation. A digital in-line vaporizer and a second CO2 absorber circumvent both of these limitations. We hypothesized that the combination of a VA reflector with an in-line vaporizer would yield substantial VA conservation, independent of fresh gas flow (FGF) in a circle circuit, and provide fine control of inspired VA concentrations. METHOD: Prospective observational study on six Yorkshire pigs. A secondary anaesthetic circuit consisting of a Y-piece with 2 one-way valves, an in-line vaporizer and a CO2 absorber in the inspiratory limb was connected to the patient's side of the VA reflector. The other side was connected to the Y-piece of a circle anaesthetic circuit. In six pigs, an inspired concentration of sevoflurane of 2.5% was maintained by the in-line vaporizer. We measured VAC at FGF of 1, 4 and 10 l/min. RESULTS: With the secondary circuit, VAC was 55% less than with the circle system alone at FGF 1 l/min, and independent of FGF over the range of 1-10 l/min. Insertion of a CO2 absorber in the secondary circuit reduced Pet CO2 by 1.3-2.0 kpa (10-15 mmHg). CONCLUSION: A secondary circuit with reflector and in-line vaporizer provides highly efficient anaesthetic delivery, independent of FGF. A second CO2 absorber was necessary to scavenge the CO2 reflected by the anaesthetic reflector. This secondary circuit may turn any open circuit ventilator into an anaesthetic delivery unit.

Technology III: in-line vaporizer with reflector.

As the clinical advantages of vapor anesthesia (VA) for sedation of patients in ICU become more apparent, the ergonomics, economy and safety issues need to be better addressed. Here we describe the use of a new commercial digital in-line anesthetic vaporizer that can be attached to the inspiratory limb of a ventilator. If used with a simple, and easily assembled secondary circuit and anesthetic reflector, the circuit remains remote from the patient, the VA consumption approaches a physical minimum, VA level is controlled and monitored, and the tidal volume size is not limited.

Effectiveness of platelet inhibition on major adverse cardiac events in non-cardiac surgery after percutaneous coronary intervention: a prospective cohort study.

BACKGROUND: Platelet inhibition is mandatory therapy after percutaneous coronary intervention (PCI). Withdrawal of oral antiplatelet agents has been linked to increased incidence of postoperative adverse cardiac events in post-PCI patients having non-cardiac surgery (NCS). There is limited knowledge of temporal changes in platelet inhibition in this high-risk surgical population. We therefore performed a multicentre prospective cohort study evaluating perioperative platelet function and its association with postoperative major adverse cardiac events (MACE). METHODS: In 201 post-PCI patients having NCS, we assessed the association between platelet function and postoperative MACE. We performed perioperative platelet function testing using a platelet mapping assay (PMA). Troponin-I was measured every 8 h for 2 days, then daily until day 5. Myocardial infarction was assessed using the third universal definition. We used multivariable logistic regression to assess the association between platelet inhibition and MACE. RESULTS: Major adverse cardiac events occurred in 40 patients within 30 days of surgery. Thirty-two of these events were non-ST-elevation myocardial infarction, four ST-elevation myocardial infarction, and four exacerbation of congestive heart failure. We were unable to show an association between platelet inhibition and MACE. The PMA showed declining levels of platelet inhibition the longer the antiplatelet therapy was withheld before surgery. Logistic regression did not show an association between preoperative platelet function or the type of stent and MACE. We found an increased cardiac risk of MACE after surgery within 6 weeks of PCI. CONCLUSIONS: The incidence of MACE in patients undergoing NCS after previous PCI is high in spite of adequate perioperative antiplatelet therapy. CLINICAL TRIAL REGISTRATION: NCT 01707459 (registered at http://www.clinicaltrials.gov).

The role of point-of-care platelet function testing in predicting postoperative bleeding following cardiac surgery: a systematic review and meta-analysis.

This systematic review and meta-analysis appraises the utility of point-of-care platelet function tests for predicting blood loss and transfusion requirements in cardiac surgical patients, and analyses whether their use within a transfusion management algorithm is associated with improved patient outcomes. We included 30 observational studies incorporating 3044 patients in the qualitative assessment, and nine randomised controlled trials including 1057 patients in the meta-analysis. Platelet function tests demonstrated significant variability in their ability to predict blood loss and transfusion requirements. Their use within a blood transfusion algorithm demonstrated a reduction in blood loss at longest follow-up (mean difference -102.9 ml (95% CI -149.9 to -56.1 ml), p < 0.001), and transfusion of packed red cells (RR 0.86 (95% CI 0.78-0.94), p = 0.001) and fresh frozen plasma (RR 0.42 (95% CI 0.30-0.59), p < 0.001). Viscoelastic methods used in combination with other platelet function tests achieved greater reduction in blood loss (mean difference -111.8 ml (95% CI -174.9 to -49.1 ml), p = 0.0005) compared with their use alone (mean difference -90.6 ml (95% CI 166.1-15.0 ml), p = 0.02). We conclude that incorporation of point-of-care platelet function tests into transfusion management algorithms is associated with a reduction in blood loss and transfusion requirements in cardiac surgery patients.

The association between tranexamic acid and convulsive seizures after cardiac surgery: a multivariate analysis in 11 529 patients.

Because of a lack of contemporary data regarding seizures after cardiac surgery, we undertook a retrospective analysis of prospectively collected data from 11 529 patients in whom cardiopulmonary bypass was used from January 2004 to December 2010. A convulsive seizure was defined as a transient episode of disturbed brain function characterised by abnormal involuntary motor movements. Multivariate regression analysis was performed to identify independent predictors of postoperative seizures. A total of 100 (0.9%) patients developed postoperative convulsive seizures. Generalised and focal seizures were identified in 68 and 32 patients, respectively. The median (IQR [range]) time after surgery when the seizure occurred was 7 (6-12 [1-216]) h and 8 (6-11 [4-18]) h, respectively. Epileptiform findings on electroencephalography were seen in 19 patients. Independent predictors of postoperative seizures included age, female sex, redo cardiac surgery, calcification of ascending aorta, congestive heart failure, deep hypothermic circulatory arrest, duration of aortic cross-clamp and tranexamic acid. When tested in a multivariate regression analysis, tranexamic acid was a strong independent predictor of seizures (OR 14.3, 95% CI 5.5-36.7; p < 0.001). Patients with convulsive seizures had 2.5 times higher in-hospital mortality rates and twice the length of hospital stay compared with patients without convulsive seizures. Mean (IQR [range]) length of stay in the intensive care unit was 115 (49-228 [32-481]) h in patients with convulsive seizures compared with 26 (22-69 [14-1080]) h in patients without seizures (p < 0.001). Convulsive seizures are a serious postoperative complication after cardiac surgery. As tranexamic acid is the only modifiable factor, its administration, particularly in doses exceeding 80 mg.kg(-1), should be weighed against the risk of postoperative seizures.

Post-operative hypercapnia-induced hyperpnoea accelerates recovery from sevoflurane anaesthesia: a prospective randomised controlled trial.

BACKGROUND: The time to recovery from vapour anaesthesia is shortened by an increase in ventilation while maintaining normocapnia. Hypercapnia during emergence from anaesthesia in spontaneously breathing patients also increases anaesthetic clearance from the brain by increasing cerebral blood flow. We hypothesised that hypercapnia-induced hyperpnoea accelerates emergence from sevoflurane anaesthesia compared to the standard anaesthesia protocol. METHODS: After Ethics Review Board approval, 44 ASA I-III patients undergoing elective gynaecological surgery were randomised after surgery to either hypercapnic hyperpnoea or control groups. In the hypercapnic hyperpnoea group, the end-tidal CO2 was adjusted to a range of 6.0-7.3 kPa to maintain a minute ventilation of 10-15 l/min. Recovery indices were compared using unpaired t-tests and ANOVA. RESULTS: Prior to extubation, minute ventilation and end-tidal CO2 in hypercapnic hyperpnoea and control groups were 10.3 ± 1.7 l/min vs. 5.4 ± 1.2 l/min (P < 0.001) and 6.6 ± 0.6 kPa and 5.2 ± 0.5 kPa (P < 0.001), respectively. Compared to control, the study group had shorter time to extubation [4.4 ± 1.3 (SD) vs. 9.8 ± 4.4 min, P < 0.01], BIS recovery to > 75 (2.4 ± 0.9 vs. 6.1 ± 3.1 min, P < 0.01), eye opening (3.9 ± 1.6 vs. 9.8 ± 6.2 min, P < 0.01), eligibility for leaving operating room (5.1 ± 1.2 vs. 11.1 ± 4.6 min, P < 0.01), and post-anaesthesia care unit (73.9 ± 14.2 vs. 89.4 ± 22.6) CONCLUSION: Hypercapnic hyperpnoea in spontaneously breathing patients halves the time of recovery from sevoflurane-induced anaesthesia in the operating room.

Pharmacokinetics of tranexamic acid in patients undergoing cardiac surgery with use of cardiopulmonary bypass.

We conducted a study to assess pharmacokinetics of high-dose tranexamic acid for 24 h after administration of the drug in patients undergoing cardiac surgery with cardiopulmonary bypass. High-dose tranexamic acid involved a bolus of 30 mg.kg(-1) infused over 15 min followed by a 16 mg.kg(-1) .h(-1) infusion until chest closure with a 2 mg.kg(-1) load within the pump prime. Tranexamic acid followed first-order kinetics best described using a two-compartment model, with a total body clearance that approximated the glomerular filtration rate. Mean plasma tranexamic acid concentrations during the intra-operative period and in the first 6 postoperative hours were consistently higher than the suggested threshold to achieve 100% inhibition and 80% inhibition of tissue plasminogen activator. With recent studies implicating high-dose tranexamic acid as a possible aetiology of postoperative seizures following cardiac surgery, the minimum effective yet safe dose of tranexamic acid in high-risk cardiac surgery needs to be refined.

Increased lung clearance of isoflurane shortens emergence in obesity: a prospective randomized-controlled trial.

BACKGROUND: There is a concern that obesity may play a role in prolonging emergence from fat-soluble inhalational anaesthetics. We hypothesized that increased pulmonary clearance of isoflurane will shorten immediate recovery from anaesthesia and post-anaesthesia care unit (PACU) stay in obese patients. METHODS: After Ethics Review Board approval, 44 ASA I-III patients with BMI>30 kg/m(2) undergoing elective gynaecological or urological surgery were randomized after completion of surgery to either an isocapnic hyperpnoea (IH) or a conventional recovery (C) group. The anaesthesia protocol included propofol, fentanyl, morphine, rocuronium and isoflurane in air/O(2) . Groups were compared using unpaired t-test and ANOVA. RESULTS: Minute ventilation in the IH group before extubation was 22.6 ± 2.7 vs. 6.3 ± 1.8 l/min in the C group. Compared with C, the IH group had a shorter time to extubation (5.4 ± 2.7 vs. 15.8 ± 2.7 min, P<0.01), initiation of spontaneous ventilation (2.7 ± 2.3 vs. 6.5 ± 4.5 min, P<0.01), BIS recovery >75 (3.2 ± 2.3 vs. 8.9 ± 5.8 min, P<0.01), eye opening (4.6 ± 2.9 vs. 13.6 ± 7.1 min, P<0.01) and eligibility for leaving the operating room (7.1 ± 2.9 vs. 19.9 ± 11.9 min, P<0.01). There was no difference in time for eligibility for PACU discharge. CONCLUSION: Increasing alveolar ventilation enhances anaesthetic elimination and accelerates short-term recovery in obese patients.

Morphometric analysis of occipital bone in the domestic cat in comparison with selected skull size parameters and with special regard to skull morphotype.

The aim of this study was to elaborate criteria defining the morphotype and to perform a morphological and morphometric analysis of the squamous part of the occipital bone and of the foramen magnum in the European cat. The study material comprised 50 corpses of European cats of both sexes and of bodyweight from 1.35 to 7.7 kg, aged from 1 year to 17 years. The study material underwent detailed preparation and morphometries of the skull, squamous part of the occipital bone and the foramen magnum were performed. The skull index (IC) data obtained indicate that the European cat represents a mesaticephalic morphotype. In the morphometrical analysis of the foramen magnum the following were included: the foramen magnum index (IFM), the occipital index (IOF), and the index of the squamous part of the occipital bone (ISO). In the morphology of the squamous part of the occipital bone two categories were distinguished: the first was characterized by a form close to an isosceles triangle with its base directed to the bottom. In none of the specimens studied was a dorsal notch in the form of a "keyhole" observed, nor was there any other deformation in the foramen magnum, which takes the form of a slightly crosswise elongated regular oval. The results of this study indicate that in the European cat the foramen magnum is free from any pathology and its shape, in the individual development, is conservative.

The expression of PDGF, its receptors and capillary morphometry in the developing lung: quantitative studies.

The aim of the present study was the evaluation of the expression levels of PDGF-A and B, and PDGFR-alpha, andchanges of capillary cross-sectional areas during terminal vascular network formation in the developing lung. Pathogen-free fetuses and newborns used for rats were used. Lungs were processed for frozen section immunohistochemical staining and parallel tissue specimens were prepared for transmission electron microscopy. The expression levels of PDGF-A and B, and PDGFR-alpha were obtained using quantitative image analysis. Additionally. the morphometry of capillary cross-sectional area was performed. We found that the expression levels of the studied antigens fluctuated, and the highest values were found in21-day old fetuses (p < 0.0001). Although there were no significant differences of the area of "a single" capillary, the area of the endothelium decreased significantly in perinatal period (p <0.0001). We conclude that the statistically significant changes of the studied antigens' expression as well as the of capillary cross-sectional area occur in the perinatal period,

Differential expression of GAD(65) and GAD(67) during the development of the rat retina.

The development of synthetic enzymes in the GABAergic system (GAD(67) and GAD(65)) of the rat retina was analyzed from birth to the 4th postnatal week by the reverse transcriptase polymerase chain reaction (RT-PCR) and by immunohistochemistry. As previously observed for GABA, immunoreactive GAD(67) profiles are seen clearly in the inner retinal layers at birth. At the end of the 1st week of postnatal life, immunolabeling is detected in amacrine and/or ganglion cells and in horizontal cells. GAD(67) immunoreactivity is transiently expressed in horizontal cells and disappears during the 3rd postnatal week. GAD(65) however does not develop until the 5th postnatal day. Immunolabeling is detected in the processes layering the inner plexiform layer (IPL) before being detected in the amacrine and/or ganglion cell bodies. The appearance of transcripts for GAD coincided with the appearance of the proteins. A transient form of mRNA transcripts of the GAD(67) gene containing an extra exon (ES-exon) is also observed which disappears progressively from birth to the 4th postnatal week. This form synthesizes a truncated, enzymatically inactive protein, which could participate in the regulation of GABA synthesis from glutamate present at high levels during retinogenesis.

Quantitative morphometry--an easy and inexpensive method for comparative studies of antigen expression during development.

The morphometric studies due to the development of computerized techniques became a very suitable tool for quantitative analysis. In the present paper we describe an easy and relatively inexpensive method useful for comparative studies on changes of expression of examined molecules during development of tissue. We used with a success this method for studies on the expression of CD31 adhesion molecule during the fetal and newborn lung development. We found that expression of CD31 is regulated during the perinatal period. It might be related to the dynamic changes in developing vascular bed in rat lung.

Retinal TUNEL-positive cells and high glutamate levels in vitreous humor of mutant quail with a glaucoma-like disorder.

PURPOSE: To investigate whether retinal cell death observed in an avian glaucoma-like disorder occurs by apoptosis and whether an increase in excitotoxic amino acid concentration in the vitreous humor is associated temporally with cell death in the retina. METHODS: Presumptive retinal apoptotic nuclei were identified by histochemical detection of DNA fragmentation (by TdT-dUTP terminal nick-end labeling [TUNEL]), and vitreal concentrations of glutamate and several other amino acids were determined by high-pressure liquid chromatography with fluorometric detection in the al mutant quail (Coturnix coturnix japonica) in which a glaucoma-like disorder develops spontaneously. RESULTS: TUNEL-labeled nuclei were located mostly in the ganglion cell layer (GCL) in the retina of mutant quails 3 months after hatching. However, labeled nuclei were also observed in the inner and outer nuclear layers. At 7 months, most TUNEL-positive nuclei were detected in the inner nuclear layer, whereas labeled cells in the GCL were reduced in number. No TUNEL-labeled nuclei were detected in the retina of control quails at any age. Vitreal concentrations of glutamate and aspartate were significantly increased in 1-month-old mutant quails compared with control animals. Concentrations decreased at 3 months, and no significant differences were observed between strains at 7 months. CONCLUSIONS: Presumptive apoptotic cell death is detected from 3 months after hatching in mutant quails and is not restricted to retinal ganglion cells. Cell death appears just after a significant increase in excitotoxic amino acid concentrations in the vitreous humor, suggesting a correlation between both events.

Genotoxicity of inhalation anesthetics halothane and isoflurane in human lymphocytes studied in vitro using the comet assay.

The alkaline single cell gel electrophoresis (comet) assay was applied to study genotoxic properties of two inhalation anesthetics-halothane and isoflurane-in human peripheral blood lymphocytes (PBL). The cells were exposed in vitro to either halothane (2-bromo-2-chloro-1,1,1-trifluoroethane) or isoflurane (1-chloro-2,2,2-trifluoroethyl difluoromethyl ether) at concentrations 0.1-10 mM in DMSO. The anesthetics-induced DNA strand breaks as well as alkali-labile sites were measured as total comet length (i.e., increase of a DNA migration). Both analysed drugs were capable of increasing DNA migration in a dose-dependent manner. In experiments conducted at two different electrophoretic conditions (0. 56 and 0.78 V/cm), halothane was able to increase DNA migration to a higher extent than isoflurane. The comet assay detects DNA strand breaks induced directly by genotoxic agents as well as DNA degradation due to cell death. For this reason a contribution of toxicity in the observed effects was examined. We tested whether the exposed PBL were able to repair halothane- and isoflurane-induced DNA damage. The treated cells were incubated in a drug-free medium at 37 degrees C for 120 min to allow processing of the induced DNA damage. PBL exposed to isoflurane at 1 mM were able to complete repair within 60 min whereas for halothane a similar result was obtained at a concentration lower by one order of magnitude: the cells exposed to halothane at 1 mM removed the damage within 120 min only partly. We conclude that the increase of DNA migration induced in PBL by isoflurane at 1 mM and by halothane at 0.1 mM was not a result of cell death-associated DNA degradation but was caused by genotoxic action of the drugs. The DNA damage detected after the exposure to halothane at 1 mM was in part a result of DNA fragmentation due to cell death.

An investigation of astroglial morphology in torpedo and scyliorhinus.

The distribution and morphology of GFAP-immunoreactive cells was investigated in two elasmobranch species, Scyliorhinus canicula and Torpedo marmorata, in an attempt to distinguish between Horstmann's (1954) hypothesis that the presence of cells resembling mammalian astrocytes is a function of the thickness of the ventricular walls, and Cajal's (1911) hypothesis that astrocytes are a phylogenetic novelty found only in birds and mammals. Two types of GFAP-reactive elements were observed, but the distribution of these differed markedly between the two species. In Scyliorhinus, radial glial cells were predominant and astrocytes relatively rare. In Torpedo, on the other hand, a species in which the ventricles are atrophied and the ventricular walls extremely thick, the overwhelming majority of GFAP-labelled structures strongly resembled astrocytes; occasionally, GFAP-positive cells were observed in the ependyma of the spinal cord. These findings, together with previous results obtained by others in hagfish, provide strong evidence in favour of Horstmann's hypothesis.

Ultrastructural changes in the lung alveoli after cardiac surgical operations with the use of cardiopulmonary bypass (CPB).

Cardiac surgical procedures with the use of cardiopulmonary bypass (CPB) are commonly complicated by pulmonary dysfunction. The mechanisms of such injury are not well understood. The aim of the present study is to analyze morphologically (mainly ultrastructurally) alveolar injury, which occurred during cardiac surgical operations involving CPB, equipped with a hollow fiber oxygenator. Our study included 20 patients, aged 45-72, who underwent coronary artery bypass grafting. Lung biopsies were taken from the left upper lobe 20 minutes after stopping CPB. Pre-CPB biopsies served as controls. Tissue specimens used for electron microscopy were processed according to standard procedures. Light microscopy revealed only a few alterations in the terminal part of the respiratory tract. Frank edema was seen in some of the alveoli. Extravasated erythrocytes as well as some neutrophils were present in the alveoli and several alveolar capillaries were congested. Ultrastructural observations confirmed the above mentioned changes. Moreover, in many alveoli, extensive injury to air-blood barrier was observed. Type I pneumocytes and endothelial cells appeared swollen or necrotically changed. The cytoplasm of type II pneumocytes was swollen. In many alveoli, pulmonary surfactant could not distribute over the alveolar surface because of edema. Structures of pulmonary surfactant were also seen in alveolar capillaries. The results of this investigation suggest that CPB is associated with some injury to lung tissue. However, this injury seems to be temporary since all examined patients had an uneventful post-operative course.

Development of the fetal and newborn lung--morphometric studies (rat model).

The aim of this study was to investigate the development of the fetal lung by morphometric means. Lung specimens from 16-20 day-old fetuses, and 1, 3, 5, 14 and 21 day-old newborns were used. Tissue specimens were evaluated by light microscopy and transmission electron microscopy. The material was divided into four groups--related to four main stages of lung development (pseudoglandular, tubular, saccular and alveolar). For each developmental stage, 30 photographs of semi-thin sections were saved on computer hard disk for future morphometric measurements. OPTIMAS 4.02 software was used for evaluation of the relative area occupied by vascular, epithelial, stromal and fluid/gaseous compartments. GraphPad InStat software was used for statistical analysis. All measurements and calculations were done per observation field. The area occupied by vessels was greatest at the pseudoglandular stage (11.7%), and smallest at alveolar stage (5.7%). The stroma also occupied the largest area at the pseudoglandular stage (58.4%) and the smallest at alveolar stage (17.0%). The area occupied by epithelial cells was most extensive at the tubular stage (52.5%) and smallest at the alveolar stage (6.1%). The fluid/gas compartment was largest at the alveolar stage (70.5%) and smallest at the pseudoglandular stage (0.58%). The present results indicate that the most dynamic lung development occurs during thin air-blood barrier formation (19th day of intrauterine life to 5th day after delivery--saccular and the beginning of alveolar stage).

Size, diffusibility and transfection performance of linear PEI/DNA complexes in the mouse central nervous system.

Currently in vivo gene delivery by synthetic vectors is hindered by the limited diffusibility of complexes in extra-cellular fluids and matrices. Here we show that certain formulations of plasmid DNA with linear polyethylenimine (22 kDa PEI, ExGene 500) can produce complexes that are sufficiently small and stable in physiological fluids so as to provide high diffusibility. When plasmid DNA was formulated with 22 kDa PEI in 5% glucose, it produced a homogeneous population of complexes with mean diameters ranging from 30 to 100 nm according to the amount of PEI used. In contrast, formulation in physiological saline produced complexes an order of magnitude greater (> or = 1 micron). Intraventricular injection of complexes formulated in glu-cose showed the complexes to be highly diffusible in the cerebrospinal fluid of newborn and adult mice, diffusing from a single site of injection throughout the entire brain ventricular spaces. Transfection efficiency was followed by histochemistry of beta-galactosidase activity and double immunocytochemistry was used to identify the cells transfected. Transgene expression was found in both neurons and glia adjacent to ventricular spaces. Thus, this method of formulation is promising for in vivo work and may well be adaptable to other vectors and physiological models.

Ultrastructural studies on selected elements of the extracellular matrix in the developing rat lung alveolus.

As the respiratory system adapts to the extrauterine life, the extracellular matrix (ECM) plays an important structural and regulatory role during its development. Therefore the purpose of this investigation was to analyze ultrastructurally several elements of extracellular connective tissue during the process of rat lung development. Morphological observations were mainly focused on the terminal part of respiratory system. To outline different components of connective tissue network, several ultrastructural techniques were used (both histochemical and immunohistochemical). The distribution and amount of the following proteins were studied: laminin, collagen type IV, collagen fibrils (CFs), elastic fibers (EFs) and fibronectin (FN). Additionally localization of glycosaminoglycans (GAGs) was examined. The present study deals with four periods of lung development: pseudoglandular, canalicular, saccular and alveolar. In all these stages localization and amount of ECM components change rapidly. In early periods of lung development, the amount of connective tissue fibers was low, basement membranes (BMs) were incomplete, and FN was distributed nearly uniformly. Later when the process of lung alveoli formation begins, the number and thickness of both CFs and EFs rapidly increased, BMs became complete, the content and distribution of FN were irregular. In all stages of lung development GAGs were distributed in BMs and among connective tissue fibers. The results described in the present study summarize morphologically ECM changes occurring during formation of lung alveolus.