Freezing Episodes Research Articles

The necrotizing effect of pulse cryocycling on liver tissue.

Repeated cycles of freezing improve the necrotizing effect of cryosurgery. We investigated whether multiple, very short periods of freezing and thawing (pulse cryocycling) enlarged the area of cell cryonecrosis within the iceball, compared with the standard method of cryocycling. Liver cryonecrosis was produced in 3 groups of rabbits by means of 2 cycles of 5-minute freezing processes, each followed by 5 minutes of spontaneous thawing. In the control group (group 1), the freezing periods were uninterrupted. In the pulse cryocycling groups, the freezing periods consisted of repeated episodes of freezing and active thawing of 15 seconds (group 2) or 30 seconds (group 3) each. The edges of all lesions were visually marked. The correlation between marking and borders of the cryolesion were examined ultrasonographically. All animals were killed on the following day, and the liver was harvested and examined histologically. Animal experimental laboratory. Complete liver cell demise was observed up to the edge of the cryolesions in all 3 groups of animals. However, a thin, sharply bounded intermediate zone of incomplete tissue destruction was observed at the border of the cryolesions, which was relatively thicker in group 2. In our study, pulse cryocycling had no advantage compared with regular cryocycling, which obtained optimal results in normal liver tissue. However, compared with the 30-second cycles, the 15-second pulse cycling yielded poorer results.

Nitrogen Dynamics in Ice Storm-Damaged Forest Ecosystems: Implications for Nitrogen Limitation Theory

Despite the widely recognized importance of disturbance in accelerating the loss of elements from land, there have been few empirical studies of the effects of natural disturbances on nitrogen (N) dynamics in forest ecosystems. We were provided the unusual opportunity for such study, partly because the intensively monitored watersheds at the Hubbard Brook Experimental Forest (HBEF), New Hampshire, experienced severe canopy damage following an ice storm. Here we report the effects of this disturbance on internal N cycling and loss for watershed 1 (W1) and watershed 6 (W6) at the HBEF and patterns of N loss from nine other severely damaged watersheds across the southern White Mountains. This approach allowed us to test one component of N limitation theory, which suggests that N losses accompanying natural disturbances can lead to the maintenance of N limitation in temperate zone forest ecosystems. Prior to the ice storm, fluxes of nitrate (NO3−) at the base of W1 and W6 were similar and were much lower than N inputs in atmospheric deposition. Following the ice storm, drainage water NO3− concentrations increased to levels that were seven to ten times greater than predisturbance values. We observed no significant differences in N mineralization, nitrification, or denitrification between damaged and undamaged areas in the HBEF watersheds, however. This result suggests that elevated NO3- concentrations were not necessarily due to accelerated rates of N cycling by soil microbes but likely resulted from decreased plant uptake of NO3-. At the regional scale, we observed high variability in the magnitude of NO3- losses: while six of the surveyed watersheds showed accelerated rates of NO3− loss, three did not. Moreover, in contrast to the strong linear relationship between NO3− loss and crown damage within HBEF watersheds [r2: (W1 = 0.91, W6 = 0.85)], stream water NO3− concentrations were weakly related to crown damage (r2 = 0.17) across our regional sites. The efflux of NO3− associated with the ice storm was slightly higher than values reported for soil freezing and insect defoliation episodes, but was approximately two to ten times lower than NO3− fluxes associated with forest harvesting. Because over one half of the entire year’s worth of N deposition was lost following the ice storm, we conclude that catastrophic disturbances contribute synergistically to the maintenance of N limitation and widely observed delays of N saturation in northern, temperate zone forest ecosystems.

Current preclinical findings on substances against Parkinson's disease

Striatal dopamine loss provides the premise for dopamine substitution in palliative therapy for Parkinson's disease (PD). This includes firstly L-dopa (L-3,4-dihydroxyphenylalanine, levodopa) and dopamine receptor agonists. Although this therapy has been demonstrated to induce marked clinical improvement in the early stages of PD, its use is limited in the long term by a loss of efficacy. In addition, most patients develop the "long-term L-dopa syndrome," which is characterized by a decrease in the control of Parkinsonian symptoms (decrease in the drug's effect) and the appearance of certain motor disturbances including episodes of akinetic freezing, debilitating dyskinesias, and on-off periods. The aim in developing new drugs is to achieve better therapeutic approaches. In the case of PD, the main strategies are to develop (1) alternatives to L-dopa therapy for alleviating the striatal dopamine deficit while avoiding or retarding the long-term L-dopa syndrome, (2) antidyskinetic approaches, and (3) neuroprotective drugs aimed at causal treatment of PD which is able to preserve the remaining dopaminergic neurones and to halt or at least retard the disease process. This article reviews new approaches for the treatment of PD and presents findings from our studies using a new experimental in vivo model of PD.

Importance of preanalytical handling of samples for measurement of cardiac troponin T in coronary effluent from isolated rat hearts

The isolated, buffer-perfused heart is probably the most widely used model in experimental heart research, and the coronary effluent is often analysed for markers of myocardial injury. Adsorption to surrounding materials may be a serious problem of protein measurements in solutions with low protein concentrations. The aims of the present study were to investigate the importance of the preanalytical phase when measuring cardiac troponin T (cTnT) in a buffer perfusate and to investigate whether addition of albumin to the effluent might increase recovery of cTnT and improve the assay. Coronary effluent was collected in tubes of different materials and in tubes with 40 g/L bovine albumin, and then frozen. cTnT was analysed at different time points after withdrawal from the freezer. cTnT was 2.3-119 times higher in effluent with albumin. In effluent without albumin, cTnT concentration declined to 2% of the initial concentration after two episodes of freezing and thawing. The cTnT loss could not be prevented by using polystyrene or siliconized glass, but was partially inhibited in effluent with albumin. Furthermore, creatine kinase and lactate dehydrogenase levels were higher in effluent with albumin. The within-series coefficient of variation for cTnT was markedly improved when using effluent with albumin.

Freeze-Thaw Effects on Metabolic Enzymes in Wood Frog Organs

To determine whether episodes of natural freezing and thawing altered the metabolic makeup of wood frog (Rana sylvatica) organs, the maximal activities of 28 enzymes of intermediary metabolism were assessed in six organs (brain, heart, kidney, liver, skeletal muscle, gut) of control (5°C acclimated), frozen (24 h at −3°C), and thawed (24 h back at 5°C) frogs. The enzymes assessed represented pathways including glycolysis, gluconeo-genesis, amino acid metabolism, fatty acid metabolism, the TCA cycle, and adenylate metabolism. Organ-specific responses seen included (a) the number of enzymes affected by freeze-thaw (1 in gut ranging to 17 in heart), (b) the magnitude and direction of response (most often enzyme activities decreased during freezing and rebounded with thawing but, liver showed freeze-specific increases in several enzymes), and (c) the response to freezing versus thawing (enzyme activities in gut and kidney changed during freezing, whereas most enzymes in skeletal muscle responded to thawing). Overall, the data show that freeze-thaw implements selected changes to the maximal activities of various enzymes of intermediary metabolism and that these may aid organ-specific responses that alter fuel use during freeze-thaw, support cryoprotectant metabolism, and aid organ endurance of freeze-induced ischemia.

Aufeis in the Ivishak River, Alaska, mapped from satellite radar interferometry

Aufeis deposits form every year on many rivers on the North Slope of Alaska; and, through repeated episodes of overflow and freezing, they may reach a thickness of more than 3 m. The presence of aufeis in a drainage basin tends to stabilize river discharge in the same way as a glacier does, by providing melt water during hot dry periods. This distribution of aufeis deposits along the rivers is discontinuous because of variations in channel geometry and under-ice water supply, and monitoring their growth during the winter is generally impractical. However, experiments using satellite radar interferometry (SRI) to map changes in the extent of aufeis near the junction of the Ivishak and the Echooka Rivers in the northern Brooks Range during the winter have produced promising results. Interferograms of the area were made from pairs of images acquired by the European Space Agency's First Earth Remote Sensing Satellite in January through March 1994. The results indicate that aufeis deposits can be mapped on interferograms as areas in river valleys in which the radar phases are poorly correlated and radar backscatter values frequently change. The ability of SRI in monitoring subtle winter aufeis processes fills a major gap in the study of aufeis by remote sensing.

Spray Irrigation Effects on Surface-Layer Stability in an Experimental Citrus Orchard during Winter Freezes

Observations taken by two surface radiation and energy budget stations deployed in the University of Florida/ Institute for Food and Agricultural Service experimental citrus orchard in Gainesville, Florida, have been analyzed to identify the effects of sprayer irrigation on thermal stability and circulation processes within the orchard during three 1992 winter freeze episodes. Lapse rates of temperature observed from a micrometeorological tower near the center of the orchard were also recorded during periods of irrigation for incorporation into the analysis. Comparisons of the near-surface temperature lapse rates observed with the two energy budget stations show consistency between the two sites and with the tower-based lapse rates taken over a vertical layer from 1.5 to 15 m above ground level. A theoretical framework was developed that demonstrates that turbulent-scale processes originating within the canopy, driven by latent heat release associated with condensation and freezing processes from water vapor and liquid water released from sprayer nozzles, can destabilize lapse rates and promote warm air mixing above the orchard canopy. The orchard data were then analyzed in the context of the theory for evidence of local overturning and displacement of surface-layer air, with warmer air from aloft driven by locally buoyant plumes generated by water vapor injected into the orchard during the irrigation periods. It was found that surface-layer lapse rates were lower during irrigation periods than under similar conditions when irrigation was not occurring, indicating a greater degree of vertical mixing of surface-layer air with air from above treetops, as a result of local convective overturning induced by the condensation heating of water vapor released at the nozzles of the sprinklers. This provides an additional explanation to the well-accepted heat of fusion release effect, of how undertree irrigation of a citrus orchard during a freeze period helps protect crops against frost damage.

RESERVOIR POTENTIAL OF GLACIO-FLUVIAL SANDSTONES: MERRIMELIA FORMATION, COOPER BASIN, SOUTH AUSTRALIA

Sedimentological analysis of cored sections within the Merrimelia Formation (basal Gidgealpa Group, Cooper Basin, S.A.) reveals a complex glacigenic environment, including glacio-lacustrine, deltaic, shorezone, fluvial, aeolian and other terrestrial facies. These facies are observed within terminoglacial and proglacial environments, and interfinger constantly throughout the Merrimelia Formation, exhibiting rapid environment change related to position of the ice sheet. Detailed sedimentological and petrological analysis, suggests that Tirrawarra sandstone-type facies belongs within the Merrimelia depositional realm. Provenance data indicates that the lithic component of the Tirrawarra Sandstone, is sourced from reworked Merrimelia coeval depositional facies. A high proportion of labile grains, which are commonly observed in the Merrimelia Formation, are only observed rarely in the Tirrawarra Sandstone (sensu stricto). It is proposed that the fluvial sandstones of the Merrimelia Formation are part of the same fluvial deposystem as the Tirrawarra Sandstone (sensu stricto) and the two sandstone sequences are the natural progression of coarsening sediment as more detritus was released from melting glaciers. It is suggested that there no longer remains any sedimentological, stratigraphical or petrographical reasons why the Tirrawarra Sandstone should not be included within the Merrimelia Formation. It is further proposed that all the braided glacio- fluvial sandstones within the Merrimelia Formation and Tirrawarra Sandstone (sensu stricto) be grouped together as 'Tirrawarra-type' facies within the Merrimelia glacigenic domain.The concept that the glacio-fluvial sandstones within the Merrimelia Formation are of the same genetic origin as the Tirrawarra Sandstone (sensu stricto), must point to the economic potential of similar sandstones found throughout the Merrimelia Formation. The thickness, sediment style and lateral extent of the proglacial braidplains is controlled by the duration of glacial melting. The Merrimelia Formation reveals a sequence of major freezing and thawing episodes with the overlying Patchawarra Formation representing the cessation of glacial influence in the Cooper Basin. Therefore, the youngest 'Tirrawarra-type' sandstone facies found at the top of the Merrimelia Formation is likely to contain the least amount of labile grains, a highly rigid siliceous framework and will be well sorted. The risk of lower reservoir quality increases down section where, with time, freezing dominated over thawing and the sediments were less rinsed, and where the resultant sandstones are more likely to be thin, more poorly sorted and chemically immature with a high proportion of labile framework grains. Thus compositional variation, controlling the diagenetic overprint, together with sandstone facies and bedform style exert profound control on the reservoir quality in the Tirrawarra-Moorari Field area, on potential gas reservoirs in the Merrimelia Formation.

Seasonal changes in plasma membrane glucose transporters enhance cryoprotectant distribution in the freeze-tolerant wood frog

One of the critical adaptations for freeze tolerance by the wood frog, Rana sylvatica, is the production of large quantities of glucose as an organ cryoprotectant during freezing exposures. Glucose export from the liver, where it is synthesized, and its uptake by other organs is dependent upon carrier-mediated transport across plasma membranes by glucose-transporter proteins. Seasonal changes in the capacity to transport glucose across plasma membranes were assessed in liver and skeletal muscle of wood frogs; summer-collected (June) frogs were compared with autumn-collected (September) cold-acclimated (5 °C for 3–4 weeks) frogs. Plasma membrane vesicles prepared from liver of autumn-collected frogs showed 6-fold higher rates of carrier-mediated glucose transport than vesicles from summer-collected frogs, maximal velocity (Vmax) values for transport being 72 ± 14 and 12.0 + 2.9 nmol∙mg protein−1∙s−1, respectively (at 10 °C). However, substrate affinity constants for carrier-mediated glucose transport (K1/2) did not change seasonally. The difference in transport rates was due to greater numbers of glucose transporters in liver plasma membranes from autumn-collected frogs. The total number of transporter sites, as determined by cytochalasin B binding, was 8.5-fold higher in autumn than in summer. Glucose transporters in wood frog liver membranes cross-reacted with antibodies to the rat GluT-2 glucose transporter (the mammalian liver isoform), and Western blots further confirmed a large increase in transporter numbers in liver membranes from autumn- versus summer-collected frogs. By contrast with the liver, however, there were no seasonal changes in glucose-transporter activity or numbers in plasma membranes isolated from skeletal muscle. We conclude that an enhanced capacity for glucose transport across liver, but not muscle, plasma membranes during autumn cold-hardening is an important adaptation that anticipates the need for rapid export of cryoprotectant from liver during natural freezing episodes.

Motor blocks in Parkinson's disease

Freezing episodes and related phenomena (as a general term, motor blocks [MBs]) are poorly understood, particularly disabling, and a therapeutically frustrating problem in Parkinson's disease (PD). Epidemiologic and clinical characteristics of MBs, as well as risk factors to develop MBs, have never been fully addressed. Herein, we report our database survey on 990 PD patients, of whom 318 (32%) had MBs. The majority of MBs were linked to gait. Start hesitation occurred in 86%, blocking on turning in 45%, and blocking in narrow spaces in 25% of patients. Initial parkinsonian symptoms in the upper body and tremor as the initial motor symptom were less likely to be associated with the presence of MBs (odds ratios [OR] 0.6 and 0.7, respectively), while initial symptoms affecting gait or trunk had higher association with MBs (OR = 1.58). Longer disease duration, higher Hoehn and Yahr stage, and longer duration of levodopa treatment are all significantly associated with the presence of MBs. We observed significant association between the existence of MBs and levodopa-induced dyskinesias to suggest similar pathophysiology. We propose that MBs in PD are abnormal retrieval or execution of complex motor tasks that can occur as a result of disease progression or as short- or long-term side effects of levodopa treatment.

Resumption of physiological functions in the wood frog (Rana sylvatica) after freezing

We monitored the resumption of physiological functions in frogs that were frozen at -2 to -3 degrees C for 24 h and thawed rapidly (at 23-25 degrees C) or slowly (at 6-8 degrees C). Bodily functions were restored sooner during fast thawing, but this did not enhance the survival of frogs. The first physiological parameter to return was cardiac function, but during the early stages of thawing heart rates were lower than heart rates of unfrozen frogs at comparable body temperatures. Heart rates increased thereafter in conjunction with the rise in frog body temperatures. Spontaneous breathing and hindleg reflexes resumed after cardiac function, but neither response was exhibited by all frogs after the conclusion of the observation periods (3-4 h). Finally, isolated gastrocnemius muscles that had undergone in vitro freezing showed no significant (P greater than 0.05) impairment of twitch and tetanic tensions even as soon as 1 h after the onset of thawing. Body systems thus vary in their rates of recovery after nonlethal freezing episodes. Furthermore, recovery of specific body systems corresponds to essential needs that must be met immediately after thawing, such as reperfusion of body tissues.

Evaluation of a modified inverted walking stick as a treatment for Parkinsonian freezing episodes

We evaluated a modified inverted walking stick [visual cue (VC) stick] in Parkinson's disease patients with freezing episodes. Patients underwent baseline trials on a 60-ft track with four walking conditions: unassisted, with a straight walking stick, with the VC stick, and on the track with parallel lines spaced every 12 in. Patients completed three trials of each condition in a randomized order, with total course time and number of freezes recorded. Patients were given the VC stick to use at home until subjective maximum functional benefit was obtained and were then brought back for a follow-up objective evaluation. Eight patients completed the study. The straight stick and VC stick worsened patient performance as a group, while the parallel lines significantly improved group performance. However, two of the eight patients showed benefit from the VC stick. Improvement with parallel lines did not predict improvement with the VC stick. On follow-up trials, the two who initially showed improvement with the VC stick continued to show improvement, while none of the other patients showed additional benefit from a learning effect. We conclude that the VC stick will help a small subpopulation of Parkinson's disease patients with freezing, and this subpopulation can be identified with the initial office visit trial.

Rapid detection of giardia antigen in stool with the use of enzyme immunoassays.

Enzyme-linked immunosorbent assays (ELISAs) using sera from rabbits and goats immunized with Giardia trophozoites were compared for detection of Giardia antigen in 300 stool specimens, 80 of which had positive results for Giardia by microscopic examination. The diagnostic accuracy of the two assays was similar, with sensitivities of 92% and 87% and specificities of 87% and 91% with the use of rabbit and goat antisera, respectively. Both ELISAs detected Giardia antigen in stool samples from asymptomatic as well as symptomatic excretors and from treated patients after organisms were no longer visualized by microscopic examination. The specificity of the assays was confirmed by consistently negative results on stool specimens from 10 neonates and 27 patients with enteric parasitic infections other than Giardia. Negative results also occurred when stool specimens containing 21 bacterial enteropathogens were tested. Potential confounding variables including multiple freezing and thawing episodes, prolonged storage, and stool filtration did not affect test results from clinical specimens. Antidiarrheal compounds did not interfere with assay results. Preservation of specimens in formalin did interfere with the assay, even if formalinized stool specimens were dialyzed before testing. For diagnosis of giardiasis, the ELISA is a sensitive and specific test that is not influenced by many environmental factors or by other enteropathogens. This test provides a practical and reliable method for evaluating large numbers of specimens in a variety of clinical and epidemiologic settings.

Chapter 25 Transplantation of fetal dopamine neurons in primate brain reverses MPTP induced parkinsonism

Parkinsonian signs were produced in the African Green monkeys by the administration of the toxin, 1-methyl-4-phenyl-l,2,3,6-tetrahydropyridine (MPTP). The transplantation of fetal neurons from the substantia nigra into two monkeys with severe symptoms of hypokinesia, postural and resting tremor, the episodes of movement freezing, and difficulty in initiating movements resulted in a reversal of these symptoms. The functional degree of recovery corresponded with the immunohistochemical observation of tyrosine hydroxylase (TH) positive neurons in the host caudate nucleus and putamen in these two animals. Dopamine metabolite levels in cerebrospinal fluid (CSF) were also elevated in these monkeys. A third monkey with parkinsonian signs received nigral tissue into a control site, the cingulate cortex. This animal failed to show recovery from the toxin-induced parkinsonism. Both early and late gestational tissue survived and showed growth of neurites, which suggested a wide developmental window in the primate brain. The combination of morphological evidence of survival and growth, the elevated neurochemical indices, and the reversal of behavioral motor impairment following the transplantation of fetal neurons into monkeys with parkinsonian signs may provide therapeutic insights into the treatment of Parkinson's disease.

Long-term study of pergolide in Parkinson's disease.

In 18 patients with Parkinson's disease, the effects of pergolide after 28 months of treatment were compared with the response after the initial 10-week therapy. At a mean 3.2-mg daily dose of pergolide, the daily dose of levodopa was still 33% lower than at the onset of pergolide therapy. The mean motor disability score, which decreased by 65% during the first 10 weeks of pergolide, was still decreased by 42% after 28 months. In the 12 patients with on-off effect, the percent time on increased 117% during the first phase of the study and was still increased 63% after more than 2 years of pergolide therapy. Sudden freezing episodes became the most disabling problem in the majority of patients. "Down-regulation" of dopamine receptors may contribute, but it is not the only cause of loss of responsiveness to pergolide.