Physiological and behavioural stress responses in cynomolgus macaques (Macaca fascicularis) to noise associated with construction work.

The aim of the present study was to evaluate the behavioural and physiological responses to environmental disturbances (live and recorded dynamite explosions) in laboratory non-human primates in preparation for a future tunnel construction underneath our animal facility. In a pilot study (A) on 20 female Macaca fascicularis, a day of test blasts resulted in an increase in faecal cortisol and immunoreactive cortisol metabolites (CICM), and the animals reacted behaviourally with vertical flight and vocalizations. In a follow-up study (B), we assessed the impact of 10 days of exposure to recorded detonations on the behaviour and CICM in 16 M. fascicularis. In the latter study we introduced a predictive signal, serving as a conditional stimulus, to half of the animals. We found no significant effects of the noise in the Signal group; while the Control groups' CICM values were affected. The behaviour was largely unaffected in the two groups. It was decided not to introduce a research moratorium on biomedical research planned to be conducted during the future tunnel construction, and that a conditional stimulus ('warning signal') will be used.

Increased (6 exon) interleukin-7 production after M. tuberculosis infection and soluble interleukin-7 receptor expression in lung tissue.

Interleukin-7 (IL-7) and the IL-7 receptor (IL-7R) have been shown to be alternatively spliced in infectious diseases. We tested IL-7 and IL-7R splicing in a tuberculosis (TB)-vaccine/Mycobacterium tuberculosis (Mtb)-challenge model in non-human primates (NHPs). Differential IL-7 splicing was detected in peripheral blood mononuclear cells (PBMCs) from 15/15 NHPs showing 6 different IL-7 spliced isoforms. This pattern did not change after infection with virulent Mtb. We demonstrated increased IL-7 (6 exon) and IL-17 protein production in lung tissue along with concomitant decreased transforming growth factor-ß (TGF-ß) from NHPs (vaccinated with a recombinant BCG (rBCG)) who showed increased survival after Mtb challenge. IL-7 increased IL-17 and interferon-γ (IFN-γ) gene and protein expression in PBMCs. Mtb-infected NHPs showed differential IL-7R splicing associated with the anatomical location and tissue origin, that is, in lung tissue, hilus, axillary lymph nodes (LNs) and spleen. Differential splicing of the IL-7R was typical for healthy (non-Mtb infected) and for Mtb-infected lung tissue with a dominant expression of soluble IL-7R (sIL-7R) receptor lacking exon 6 (9:1 ratio of sIL-7R/cell-bound IL-7R). Differential ratios of cell-bound vs sIL-7R could be observed in hilus and axillary LNs from Mtb-infected NHPs with an inversed ratio of 1:9 (sIL-7R/cell-bound IL-7R) in spleen and PBMCs. Soluble IL-7R is exclusively present in lung tissue.

Positive reinforcement training in rhesus macaques-training progress as a result of training frequency.

Positive reinforcement training (PRT) efficiency was examined as a function of training frequency in 33 pair- or triple-housed female rhesus macaques. The animals were trained three times a week, once a day or twice a day, using PRT and a clicker as a secondary reinforcer. All animals were trained on 30 sessions, with an average of 5 min per training session per animal. The behaviors, trained in succession, were Targeting (reliably touching and following a Target); Collaborating (dominant animals allowing subordinates to train while stationing); Box-training (accepting being enclosed in a small compartment while responding to Target training) and initial Injection training.Fulfilled criteria for Targeting were obtained in 32/33 animals in a median of nine training sessions. Collaboration was obtained in 27/33 animals in a median of 15 training sessions. However, only four animals completed Box-training during the 30 training sessions and started Injection training. When comparing training success in terms of number of training sessions, training twice a day was less efficient than the other two treatments. In terms of daily progress, our results suggest that from a management perspective, daily training is more conducive to quick training success than thrice weekly training. In addition, in this study no further advantages could be gained from training twice a day.

Pathology of the cochlea following a spontaneous mutation in DBA/2 mice.

The DBA/2 strain of mice usually presents with noise-induced epileptic seizures and hearing disorders. After a spontaneous mutation a strain with early hearing loss and circling behaviour was produced. This strain presents with clinical symptoms found in diseases connected to inner ear disorders. These animals do not suffer from periodical disorders, however, but have functional disturbances continuously and can therefore serve as an animal model for diseases originating from both parts of the inner ear. The genetic inheritance appears to be autosomal recessive. Offspring showed circling behaviour and severe pathology in the vestibular part of the inner ear. In the present study pathology of the cochlear part of the inner ear was visualized using conventional microscopical techniques. The content of actin and fodrin was labelled immunohistochemically, and hearing was assessed with auditory brainstem recordings. After 1 month the animals showed deterioration of the cochlear part of the inner ear. At 6 months no organ of Corti remained and the animals were deaf. Transmission and scanning electron microscopy revealed severe apical hair cell changes. The content of alpha-actinin and fodrin in the DBA/2 mouse was already fainter than that in age-matched CBA control mice at the age of 1 month. Labelling of antibodies against fodrin increased in the supporting cells of the older animals, probably owing to the replacement of hair cells.

Morphological and functional characteristics of the different cell types in the stria vascularis: a comparison between cells obtained from fresh tissue preparations and cells cultured in vitro.

Endolymph is the only extracellular fluid in the body which is characterized by an intracellular-like ion composition. The molecular mechanisms responsible for the production of endolymph and the regulation of endolymph composition are still unknown to a large extent, although the stria vascularis (SV) is believed to play an important role for these functions. A basic requirement for investigating the function of different cell types in the SV is the establishment of a method, which increases the accessibility of the tissue with maintained cell viability and function. In this study, fresh tissue preparations and cultured cells from SV, harvested from pigmented guinea pigs, were established. Marginal cells, intermediate/melanocyte-like cells and fibroblasts could be discerned in the cell cultures with bright-field microscopy, transmission and scanning electron microscopy as well as immunohistochemistry, using polyclonal antibodies against cytokeratin and vimentin. In order to study functional characteristics of the fresh tissue preparations and the cell cultures, changes in the cytoplasmic free Ca2+ concentration were determined with the fura-2 method. The cultured cells, of different types in the SV, are a suitable model for future studies of the molecular mechanisms behind the production of endolymph and the regulation of endolymph composition.

Fibrinous reaction on implanted intraocular lenses. A comparison of conventional PMMA and heparin surface modified lenses.

We studied the fibrinous reaction after intraocular lens (IOL) implantation in the posterior chamber of cynomolgus monkeys. In 50% of the eyes, we implanted an IOL made of conventional poly-(methyl methacrylate) (PMMA); in the remaining eyes we implanted a PMMA IOL with a heparin modified surface. Two, 4, 8, and 18 weeks after surgery the eyes were examined by slitlamp for fibrinous reactions on and around the IOL surface. At weeks 4, 8, and 18 there was a marked decrease in fibrinous reaction in the eyes with a heparin surface modified IOL. The results of this study accord with earlier findings that heparin surface modification improves the biocompatibility of the IOL.

Improved biocompatibility of intraocular lenses by heparin surface modification: a 12-month implantation study in monkeys.

To evaluate the long-term biocompatibility and potential side effects of heparin surface modification of a poly(methyl methacrylate) intraocular lens (IOL), a heparin surface modified IOL was implanted in the left posterior chamber of 24 cynomolgus monkeys and a reference IOL (without surface modification) was implanted in the right eye in 12 of these animals. Twelve eyes were not operated on. Eleven eyes in seven monkeys were lens extracted as a control of the surgical method. Slitlamp examinations and intraocular pressure recordings were made one day, one and two weeks, and 1, 2, 2 1/2, 3 1/2, 6, 8, 10, and 12 months after the operation. Eleven monkeys were sacrificed after 3 1/2 months and the remaining animals after 12 months for morphological examination of the eyes. Slitlamp and morphological examinations showed that cell deposits, pigmentation, and posterior synechias were significantly less in eyes with heparin surface modified IOLs than in eyes with reference IOLs throughout the 12-month observation period. The intraocular pressure was equally reduced in eyes with heparin surface modified IOLs and reference IOLs for about one month, after which it returned to normal. No side effects following the implantation of heparin surface modified IOLs were observed. We concluded that heparin surface modification of IOLs is efficient for long-term reduction of cell deposits and posterior synechias after implantation in monkey eyes and may also be effective in lowering the degree of side effects to IOL implantation in humans.

Does ciprofloxacin affect the inner ear? A preliminary report.

Ciprofloxacin, a 4-quinolone antibiotic was tested regarding its possible influence on the inner ear sensory epithelia. Adult guinea pigs were injected intraperitoneally with ciprofloxacin in a dosage of 25, 50, 100 and 150 mg/kg body weight per day. The animals in the high dosage ranges stopped eating and consequently lost weight quite rapidly. After sacrifice, the inner ear sensory epithelia were removed for ultrastructural analysis. The general outline of the vestibular sensory epithelia as well as the cochleae were normal. Mild changes in the sensory hairs of the third row of cochlear outer hair cells were noted in the specimens obtained from the high dose animals. These changes were discrete and did not resemble any previously known pattern of ototoxic damage. At present no data indicate that ciprofloxacin has a toxic effect on the inner ear.

Adenylate cyclase activity in the fetal and the early postnatal inner ear of the mouse.

The adenylate cyclase activity was analyzed in fetal, early postnatal and adult inner ears of the CBA/CBA mouse and also in approximately one month old inner ears from Shaker -1 and Shaker -2 mice. A comparison was made with the maturation of potassium levels in endolymph as investigated with the X-ray energy dispersive technique. Adenylate cyclase activity in the developing normal inner ear shows two significant periods of increases: from the 16th to the 19th gestational day in both the cochlear and vestibular parts of the labyrinth, and from birth to day 6 after birth in the lateral wall tissues of the scala media. During the first period the anatomical boundaries of the secretory epithelia are developing. The postnatal rise in adenylate cyclase activity correlates with the morphological maturation of stria vascularis at the cellular and subcellular levels and the rise in potassium content of endolymph. The rise of enzyme activity in the cochlear during the maturation of endolymph supports a link between adenylate cyclase and the control of inner ear fluids. Adenylate cyclase activity in stria vascularis/spiral ligament of Shaker -1 and Shaker -2 mice were at normal levels and correlated better with the rather normal morphology of the tissues than the abnormal composition of endolymph in these mutants.

Biochemical studies on the embryonic development of the mammalian inner ear in organ culture.

Adenylate cyclase activity and phospholipid labeling were compared during embryonic development of the mouse inner ear in vivo and in vitro. Inner ears were explanted on the 16th gestational day and cultured in vitro for 3-12 days. The gestation time in vivo is 21 days. During the 1st week in vitro there is very little growth of the inner ear with regard to total protein content. In contrast, the labyrinth increases its protein content threefold during the corresponding period of time in vivo. The activity of adenylate cyclase develops parallel in vivo and in vitro until the 19th gestational day whereafter the specific activity of the enzyme in vitro surpasses that of the enzyme in vivo three- to fivefold suggesting a lack of control mechanisms in organ culture. Phospholipids are labeled by 32P in an essentially similar quantitative relationship in vivo and vitro, while some quantitative differences exist. According to the present study the usefulness of the organ culture for the investigation of inner ear development appears limited to a culture period corresponding to an age prior to birth.