Microstructure and thermal evolution of sea-ice brine inclusions were investigated with magnetic resonance imaging Ž . 1MRI techniques. Ice samples were kept at temperatures between y28C and y258C during H imaging in a 4.7-T magnet Ž at 200 MHz. Measurements were completed in a 20-cm diameter cylindrical probe and actively shielded gradient coils max. y1. 50 mT m , pixel dimensions ) 0.2 mm, slice thicknesses ) 1 mm , and for higher resolution in a mini-imaging unit with y1 Ž . a 9-cm diameter probe with gradient coils of 200 mT m pixel dimensions -0.1 mm, slice thickness -0.4 mm . Ž . Absorption of radio-frequency RF signals in the dielectrically lossy brine resulted in degraded signals and was alleviated Ž . Ž by use of a contrast agent decane . MRI data and sea-ice thin section images agree very well -5% deviation for pore . microstructural parameters . Analysis of ice grown under different current speeds indicates that pores are smaller and pore number densities larger at higher current speeds. The thermal evolution of fluid inclusions was studied on cold first-year ice samples, maintained at close to in-situ temperatures prior to experiments. Warming from y218C to y108C to y68C is Ž . associated with a distinct increase in pore size from 1.5 to 1.7 to 2.6 mm for the upper 10-percentile in the vertical and Ž . Ž elongation 4.0 to 4.2 to 6.2 for ratio of major to minor pore axes in the vertical and a decrease in number densities 0.75 to y3 . Ž . 0.62 to 0.58 mm in the vertical . Aspect ratios increased from 4:2:1 to 6:2:1 upper 10-percentile , indicating expansion and merging of pores in the vertical, possibly promoted by microscopic residual brine inclusions. q

In vivo magnetic resonance imaging (MRI) and angiography were applied to the marine spider crab Maja squinado for a study of temperature effects and thermal tolerance. Ventilation and haemolymph circulation were investigated during progressive cooling from 12°C to 2°C. The anatomical resolution of MR images from Maja squinado obtained with a standard spin echo sequence were suitable to resolve the structures of various internal organs. The heart of the animal could be depicted without movement artifacts. The use of a flow compensated gradient echo sequence allowed simultaneous observations of ventilation, reflected by water flow through the gill chambers as well as of haemolymph flow. Simultaneous investigation of various arteries was possible by use of flow weighted MRI. In addition to those accessible by standard invasive flow sensitive doppler sensors, flow changes in gill, leg arteries and the venous return could be observed. Both ventilation and haemolymph flow decreased during progressive cooling and changes in haemolymph flow varied between arteries. Haemolymph flow through the Arteria sternalis, some gill and leg arteries was maintained at low temperatures indicating a reduced thermal sensitivity of flow in selected vessels. In support of previous invasive studies of haemolymph flow as well as heart and ventilation rates, the results demonstrate that the operation of gills and the maintenance of locomotor activity are critical for cold tolerance. A shift in haemolymph flow between arteries likely occurs to ensure the functioning of locomotion and ventilation in the cold.

Temperature-dependent adjustments of intracellular pH are thought to play a major role in the maintenance of protein function. Comparative studies were carried out in two species from the same ®sh family (Zoarcidae), the stenothermal Antarctic eelpout (Pachycara brachycephalum) and the eurythermal eelpout (Zoarces viviparus), to ®nd out whether pH regulation is modi®ed by temperature in the closely related species and to what extent the respective pattern diers between eurytherms and stenotherms. Previous invasive studies had compared individual animals sampled at various temperatures and suggested that a decrease in intracellular pH (pHi) values occurs at rising temperatures, as predicted by the alpha-stat hypothesis of acidbase regulation. The present study used non-invasive in vivo 31 P-NMR spectroscopy in non-anaesthetized, unrestrained ®sh for long-term online recordings in individual specimens. Control spectra obtained at T=0°C for P. brachycephalum and at 12°C for Z. viviparus indicated low stress conditions, as well as a high stability of energy and acid-base status over time periods longer than 1 week. Temperature changes had no in¯uence on the concentration of high-energy phosphates like phosphocreatine or ATP. Temperature-induced pH changes were monitored continuously in a range between 0 and 6°C for polar, and 12 and 18°C for temperate eelpout. A pHi change of around ±0.015 pH units/°C was observed within both species, in accordance with the alpha-stat hypothesis; however, extrapolation to the same temperature revealed dierent set points of pH regulation in the two species. These ®ndings con®rm that an alphastat pattern of pH regulation can be found in stenothermal Antarctic animals, at set points deviating from an alpha-stat pattern, however, in a between-species comparison.

Energetic studies on exercising animals are usually limited to oxygen consumption measurements in respirometers followed by invasive tissue sampling and analysis of metabolites. Noninvasive studies of exercising animals like through the use of 31 P NMR are typically restricted to ''stop and go'' measurements. Furthermore, magnetic resonance studies of marine animals are hampered by sea water, a highly electric conductive and dielectric medium, resulting in heavy loading and strong RF loss. In this work, we present a set-up for online determination of muscle bioenergetics in swimming marine fish, Atlantic cod (Gadus morhua), using in vivo 31 P NMR spectroscopy, which overcome these limitations. Special hardware and RF coils were developed for this purpose.