Thesis (Ph.D.)--Auburn University, 2007. / Abstract. Vita. Includes bibliographic references (ℓ.71-77)

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This thesis focuses on molecular characterization of circadian clock genes in insects. It explores genetic diversity of circadian clock genes by molecular characterization of several insect species including two dipteran flies (Musca domestica and Drosophila melanogaster), two cockroach species representing ancestral insects, and the linden bug, Pyrrhocoris apterus. Furthermore it considers various roles of circadian clock genes in insect physiology. Application of molecular-biology methods in Pyrrhocoris apterus, non-model insect species, enable us to investigate involvement of circadian clock genes in photoperiod induced physiological responses. Application of molecular-biology methods in Periplaneta americana and Blattella germanica was used to explore involvement of circadian clock genes in magnetoreception.

Many animals on earth need to navigate in their own environments for breeding and foraging. The ability to determine a correct heading and the current location for a migratory bird is critical for its survival. Animals that are sensitive to the Earth’s magnetic field can use it to obtain their direction of travel. In 2000, a paper suggested that radical pair reaction could form the basis for magnetoreception in migratory birds and a flavoprotein, cryptochrome, was proposed as the candidate for the radical pair precursor. Recent in vivo experimental results strongly support the hypothesis that radical pairs formed in the eyes of migratory birds are responsible for their magnetic compass sense. Cryptochrome has also been located in the UV-cones in the retinas of two different species of bird. Radical pairs in living cells are influenced mainly by Zeeman interaction, hyperfine interaction, rotational modulation, etc., and together they influence the recombination reactions of the radical pairs. This thesis considers the possible role of radical pairs in avian magnetoreception, using computer simulations of the quantum mechanical evolution of a radical pair under a variety of conditions. Chapter 1 contains the introductions to spin chemistry, avian magnetoreception, and the mathematical description of the quantum evolution of a radical pair. Chapter 2 describes the four different theoretical models for a general non-diffusion-controlled radical pair reaction and the product yields of a radical pair reaction predicted by these four models are analysed and compared. Chapter 3 introduces a model for avian magnetoreception that integrates photoselection with the radical pair reaction and the model is used to predict the retinal patterns that a bird may be able to use for magnetoreception. The anisotropic singlet product yields of a radical pair comprises the flavin chromophore and the tryptophan of a cryptochrome are also presented in this chapter. A paper based on some parts of this chapter is published [1] in the Journal of The Royal Society Interface. Chapter 4 describes a modified version of an algorithm that is used to calculate the product yields detected in a reaction yield detected magnetic resonance (RYDMR) experiment. The new algorithm is used to analyse the results of two sets of RYDMR experiments in which two radical pair systems, pyrene/1,3-dicyanobenzene and chrysene/1,4-dicyanobenzene, were used. The modulated detection technique used in the RYDMR experiments is also discussed in this chapter.

Currently, much attention is paid to magnetoreception, a sense which allows an organism to detect a magnetic field of the earth. Now, this discovery is related to positional behaviour of animals (magnetic alignment) in the magnetic field and the world phenomenon of "magnetic cows". The magnetic alignment is a spontaneous (congenital) behaviour, when the animal puts its body axis into the positions of north - south or east - west. Most studies have demonstrated the north - south direction preference. This work contributes to clarify positional behaviour of cattle during the daily cycle. This study is based on a different data collection and is targeted at individuals in the herd. Individuals were photographed in a given direction, at hourly intervals. The magnetic alignment was determined in 26 individuals from three different cattle herds, mainly east or west. Statistically significant results during daily cycles in individual intervals also confirmed the north - south direction preference. There were also discussed the environmental conditions that may affect the final result. For inconclusive results, the individuals were probably motivated by temporarily more important local incentives.

The magnetic orientation was established in several species of rodents, but the mechanism of its perception has not been determined. Aim of this work was to verify magnetic sense in the bank vole (Myodes glareolus) and try to solve the nature of its magnetoreception in tests carried out in total darkness and when the horizontal component was reversed and the vertical component was inverted. A spontaneous directional preference in the magnetic field was tested in circular arena.

Many animals have ability to percieve the magnetic field of the Earth and use this clue for both orientation and navigation. Yet little is known about physiological mechanism that underlies this sensory ability. Although physiological mechanism still remains unclear, there are three major hypotheses how animals can detect the magnetic field. In this study I focused on the radical pair theory. I analyzed how manipulation of the ambient magnetic field influence the expression of CRY 1, CRY 2 and c-Fos in the pigeon´s retina. I observed in incerased numbers of CRY1, c-Fos and CRY1+c-Fos possitive cells in the ihned nuclear layer (INL) of the retina in animals sbjected to the periodical manipulation of the magnetic field inclination. These data demonstrate that the INL constains a population of neurons that are responsive to magnetic stimuli and strongly suggest that Cry 1 is involved in detection of the Earth magnetic field.

The mechanism behind magnetoreception – the ability to sense magnetic fields for orientation and navigation – still remains one of the most difficult questions to answer in sensory biology, with fish being just one of many taxa known to possess this sense. Characterizing a magnetic sense in fish is crucial for understanding how they navigate their environment and can inform on how increasing anthropogenic sources of electromagnetic fields in aquatic environments may affect threatened fish species. This study examined the hypothesis put forth by Natan and Vortman (2017) that magnetotactic bacteria (MTB), bacteria that create their own chains of magnetic particles for navigational use, act in symbiosis with their animal host to convey magnetic information about their surroundings. Utilizing existing, publicly available datasets of raw genomic sequences, this study demonstrated the presence of MTB within a diverse array of fishes and identified differences in species diversity of MTB between freshwater and marine species of fish. Future research aimed at identifying MTB in specific fish tissues, such as the eye and other neural tissues, will be necessary to provide support for this hypothesis and to further examine the relationships that MTB may have with magnetically sensitive animals.

Use of a magnetic compass has been demonstrated in all major classes of vertebrates as well as several classes of invertebrates, and is proposed to involve a photo-induced radical pair mechanism (RPM). My dissertation research consisted of characterizing a magnetic compass in a model species, Drosophila melanogaster. Preliminary experiments were carried out with adult flies, however, due to the behavioral complexity of adult responses a new behavioral assay of magnetic compass orientation was developed using larval Drosophila that elicits a robust magnetic compass response in a trained magnetic direction. This manuscript describes experiments that were conducted showing that larval magnetic compass orientation: 1) demonstrates a complex 3-dimensional pattern of response consistent with a RPM; 2) is consistent with a receptor mechanism that utilizes short- and long-wavelength antagonistic photopigments, proposed to explain wavelength dependent effects in vertebrates (e.g. amphibians and birds); and 3) produces axially symmetrical patterns of response with respect to the geomagnetic field. Additionally, tests of adult Drosophila under low and high intensities of monochromatic long wavelength light revealed a similar behavioral response to varying intensities of monochromatic light as previously reported in migratory birds (E. rubecula). These findings indicate that the magnetic compass of larval Drosophila shares a common functional architecture and similar biophysical mechanism with that of at least some vertebrates (e.g. amphibians and possibly birds), suggesting that the magnetic compass of modern vertebrates may have evolved once in a common ancestor of these three lineages over 450 million years ago. Furthermore, findings indicating a spontaneous preference for magnetic directions in D. melanogaster larvae suggest that a light-dependent magnetoreception mechanism is more widespread in insects than was previously suspected. The development of a behavioral assay to study the light-dependent magnetic compass in an organism with a simple nervous system, a limited behavioral repertoire, and with the possibility of using the full power of modern molecular and genetic techniques holds considerable promise to increase our understanding of the biophysical mechanism(s) and neurophysiological structures underlying magnetic orientation in terrestrial animals. / Ph. D.

Magnetoreception is the ability of some animals to use the weak magnetic field of the Earth for navigation over long-distance migrations. It is a well-known phenomenon, but its underlying biophysical mechanisms remain poorly understood. One proposal involves light-induced, magnetically sensitive chemical reactions occurring within cryptochrome proteins, rationalised via the radical pair mechanism (Chapter 1). The absence of evidence in support of this hypothesis is in part due to the lack of sufficiently sensitive techniques to measure magnetic field effects (MFEs) in biological samples. Cavity-enhanced detection, most commonly in the form of cavity ring-down spectroscopy (CRDS) or cavity-enhanced absorption spectroscopy (CEAS), is widely used in the gas phase to provide significant sensitivity gains over traditional single-pass measurements (Chapter 2). However, successful studies in the condensed phase are less prevalent due to the additional background losses inherent to the sample. This thesis reports on the application of broadband (i.e. monitoring > 100nm) variants of CRDS and CEAS to the study of MFEs on the radical recombination reactions of flavin-based systems in solution. The broadband CRDS (BBCRDS) instrument employed in Chapter 4 is able to monitor the spectral changes induced by magnetic fields with submicrosecond time resolution. However, the need to scan both the probe wavelength and time delay to construct time-resolved spectra leads to prohibitively long acquisition times, and hence exposure of sensitive samples to high numbers of photons. The broadband CEAS (BBCEAS) studies reported in Chapter 5 combine the high irradiance and spectral coverage of a supercontinuum radiation (SCR) source with a CCD detector to simultaneously acquire absorption spectra across the visible region (480–700nm). The CW nature of this technique precludes the possibility of following radical pair kinetics in real time. In an effort to combine the respective advantages of these two instruments, which individually have represented powerful advances in capability, a new cavity-enhanced technique is reported for the first time (Chapter 6). The result, optical cavity-enhanced transient absorption spectroscopy (OCTAS), is able to simultaneously monitor spectral evolution and associated MFEs on the microsecond timescale, with comparable sensitivity to the existing techniques. Magnetic responses in animal cryptochrome proteins have successfully been recorded using all three techniques, lending considerable weight to the hypothesis that these molecules are at the heart of the magnetic sense in animals.