Gene delivery to neurons in the auditory brainstem of barn owls using standard recombinant adeno-associated virus vectors.

Recombinant adeno-associated virus (rAAV) vectors are a commonly used tool for gene delivery. There is a large choice of different serotypes whose transduction efficiency varies for different animal species. In this study, three rAAV vectors were tested for transduction efficiency in the auditory brainstem of adult barn owls (Tyto alba) which are not standard laboratory animals. Injections with rAAV serotypes 2/1 and 2/5 resulted in reliable expression in various nuclei of the auditory brainstem of barn owls. Both vectors showed evidence of being spread by axonal transport. However, only rAAV2/5 also showed expression in regions far distant from the injection site, suggesting long-range axonal transport in connections along the auditory pathway. In contrast, injections with rAAV2/9 resulted in no expression. Our results demonstrate for the first time that commercially available rAAV vectors can be used for reliable gene expression in the barn owl auditory brainstem and pave the way toward optogenetic manipulation of neural activity in this important animal species in neuroethology and auditory physiology.

Gas Anesthesia Impairs Peripheral Auditory Sensitivity in Barn Owls (Tyto alba).

Auditory nerve single-unit recordings were obtained from two groups of young barn owls (age, between posthatching days 11 and 86) in terminal experiments under two different anesthetic regimes: ketamine (6-11 mg/kg) plus xylazine (∼2 mg/kg); or isoflurane (1-1.5%) in oxygen, delivered via artificial respiration. In a second series of minimally invasive experiments, auditory brainstem responses (ABRs) were recorded in the same four adult barn owls (Tyto alba; age, between 5 and 32 months) under three different anesthetic protocols: ketamine (10 mg/kg) plus xylazine (3 mg/kg), isoflurane (1-1.5%), and sevoflurane (2-3%) in carbogen. Finally, the ABR measurements on adult owls were repeated in terminal experiments including more invasive procedures such as artificial respiration and higher isoflurane dosage. The main finding was a significant deterioration of auditory sensitivity in barn owls under gas anesthesia, at the level of the auditory nerve (i.e., a very peripheral level of the auditory system). The effect was drastic in the young animals that experienced threshold elevations in auditory nerve single-unit responses of ≥20 dB. ABR thresholds assessed repeatedly in experiments on adult owls were also significantly higher under isoflurane and sevoflurane, on average by 7 and 15 dB, compared with ketamine/xylazine. This difference already occurred with minimal dosages and was reversibly enlarged with increased isoflurane concentration. Finally, there was evidence for confounding detrimental effects associated with artificial respiration over many hours, which suggested oxygen toxicity.

Migratory blackcaps can use their magnetic compass at 5 degrees inclination, but are completely random at 0 degrees inclination.

It is known that night-migratory songbirds use a magnetic compass measuring the magnetic inclination angle, i.e. the angle between the Earth's surface and the magnetic field lines, but how do such birds orient at the magnetic equator? A previous study reported that birds are completely randomly oriented in a horizontal north-south magnetic field with 0° inclination angle. This seems counter-intuitive, because birds using an inclination compass should be able to separate the north-south axis from the east-west axis, so that bimodal orientation might be expected in a horizontal field. Furthermore, little is known about how shallow inclination angles migratory birds can still use for orientation. In this study, we tested the magnetic compass orientation of night-migratory Eurasian blackcaps (Sylvia atricapilla) in magnetic fields with 5° and 0° inclination. At 5° inclination, the birds oriented as well as they did in the normal 67° inclined field in Oldenburg. In contrast, they were completely randomly oriented in the horizontal field, showing no sign of bimodality. Our results indicate that the inclination limit for the magnetic compass of the blackcap is below 5° and that these birds indeed seem completely unable to use their magnetic compass for orientation in a horizontal magnetic field.

Re-calibration of the magnetic compass in hand-raised European robins (Erithacus rubecula).

Migratory birds can use a variety of environmental cues for orientation. A primary calibration between the celestial and magnetic compasses seems to be fundamental prior to a bird's first autumn migration. Releasing hand-raised or rescued young birds back into the wild might therefore be a problem because they might not have established a functional orientation system during their first calendar year. Here, we test whether hand-raised European robins that did not develop any functional compass before or during their first autumn migration could relearn to orient if they were exposed to natural celestial cues during the subsequent winter and spring. When tested in the geomagnetic field without access to celestial cues, these birds could orient in their species-specific spring migratory direction. In contrast, control birds that were deprived of any natural celestial cues throughout remained unable to orient. Our experiments suggest that European robins are still capable of establishing a functional orientation system after their first autumn. Although the external reference remains speculative, most likely, natural celestial cues enabled our birds to calibrate their magnetic compass. Our data suggest that avian compass systems are more flexible than previously believed and have implications for the release of hand-reared migratory birds.