Loss of CNGB1 protein leads to olfactory dysfunction and subciliary cyclic nucleotide-gated channel trapping

Olfactory receptor neurons (ORNs) employ a cyclic nucleotide-gated (CNG) channel to generate a receptor current in response to an odorant-induced rise in cAMP. This channel contains three types of subunits, the principal CNGA2 subunit and two modulatory subunits (CNGA4 and CNGB1b). Here, we have analyzed the functional relevance of CNGB1 for olfaction by gene targeting in mice. Electro-olfactogram responses of CNGB1-deficient (CNGB1-/-) mice displayed a reduced maximal amplitude and decelerated onset and recovery kinetics compared with wild-type mice. In a behavioral test, CNGB1-/- mice exhibited a profoundly decreased olfactory performance. Electrophysiological recordings revealed that ORNs of CNGB1-/- mice weakly expressed a CNG current with decreased cAMP sensitivity, very rapid flicker-gating behavior and no fast modulation by Ca2+-calmodulin. Co-immunoprecipitation confirmed the presence of a CNGA2/CNGA4 channel in the olfactory epithelium of CNGB1-/- mice. This CNGA2/CNGA4 channel was targeted to the plasma membrane of olfactory knobs, but failed to be trafficked into olfactory cilia. Interestingly, we observed a similar trafficking defect in mice deficient for the CNGA4 subunit. In conclusion, these results demonstrate that CNGB1 has a dual function in vivo. First, it endows the olfactory CNG channel with a variety of biophysical properties tailored to the specific requirements of olfactory transduction. Second, together with the CNGA4 subunit, CNGB1 is needed for ciliary targeting of the olfactory CNG channel.     

Olfaction in Three Genetic and Two MPTP-Induced Parkinson’s Disease Mouse Models

Various genetic or toxin-induced mouse models are frequently used for investigation of early PD pathology. Although olfactory impairment is known to precede motor symptoms by years, it is not known whether it is caused by impairments in the brain, the olfactory epithelium, or both. In this study, we investigated the olfactory function in three genetic Parkinson’s disease (PD) mouse models and mice treated with MPTP intraperitoneally and intranasally. To investigate olfactory function, we performed electro-olfactogram recordings (EOGs) and an olfactory behavior test (cookie-finding test). We show that neither a parkin knockout mouse strain, nor intraperitoneal MPTP treated animals display any olfactory impairment in EOG recordings and the applied behavior test. We also found no difference in the responses of the olfactory epithelium to odorants in a mouse strain over-expressing doubly mutated α-synuclein, while this mouse strain was not suitable to test olfaction in a cookie-finding test as it displays a mobility impairment. A transgenic mouse expressing mutated α-synuclein in dopaminergic neurons performed equal to control animals in the cookie-finding test. Further we show that intranasal MPTP application can cause functional damage of the olfactory epithelium.     

Methane output of tortoises: its contribution to energy loss related to herbivore body mass

An increase in body mass (M) is traditionally considered advantageous for herbivores in terms of digestive efficiency. However, recently increasing methane losses with increasing M were described in mammals. To test this pattern in non-mammal herbivores, we conducted feeding trails with 24 tortoises of various species (M range 0.52-180 kg) fed a diet of grass hay ad libitum and salad. Mean daily dry matter and gross energy intake measured over 30 consecutive days scaled to M(0.75 (95%CI 0.64-0.87)) and M(0.77 (95%CI 0.66-0.88)), respectively. Methane production was measured over two consecutive days in respiration chambers and scaled to M(1.03 (95%CI 0.84-1.22)). When expressed as energy loss per gross energy intake, methane losses scaled to 0.70 (95%CI 0.47-1.05) M(0.29 (95%CI 0.14-0.45)). This scaling overlaps in its confidence intervals to that calculated for nonruminant mammals 0.79 (95%CI 0.63-0.99) M(0.15 (95%CI 0.09-0.20)), but is lower than that for ruminants. The similarity between nonruminant mammals and tortoises suggest a common evolution of the gut fauna in ectotherms and endotherms, and that the increase in energetic losses due to methane production with increasing body mass is a general allometric principle in herbivores. These findings add evidence to the view that large body size itself does not necessarily convey a digestive advantage.     

Characteristics of Acaciamangium shoot apical meristems in natural and in vitro conditions in relation to heteroblasty

Morphological and histocytological characteristics of Acacia mangium shoot apical meristems (SAMs) were assessed in natural and in vitro conditions in relation to heteroblasty. In the natural environment, SAMs with a mature-phyllode morphology were much bigger, contained more cells with larger vacuolated area, or vacuome, and lower nucleoplasmic ratios than those from the juvenile type (Juv). In these latter, nuclei appeared more voluminous, evenly and lightly stained, with clearly distinguishable nucleolei and less abundant chromocenters. In vitro, where reversions from mature to juvenile morphological traits do occur unpredictably, heteroblasty was less obvious in the SAM characteristics examined. In vitro SAMs corresponding to the juvenile and mature types showed similarities with outdoor Juv SAMs, but could be distinguished from these latter by a much larger vacuome that might be induced by the culture conditions. These findings encourage pursuing the investigations at the chromatin and nucleolus level in SAM zones where heteroblasty-related differences have been detected.     

Variation in Growth and Potentially Associated Health Status in Hermann's and Spur‐Thighed Tortoise (Testudo hermanni and Testudo graeca)

Captive reptiles often show higher growth rates than in the wild, possibly due to higher feeding intensity. Although health problems are usually linked to inappropriate diets, fast growth itself, such as triggered by appropriate diets fed in high amounts, has traditionally also been considered unfavorable for tortoises. We document growth rates (based on age and mass) from private Testudo hermanni and T. graeca breeders, which are generally higher than those reported for free‐ranging specimens, but show enormous variation. Tortoise patients presented to an exotics clinic also covered the whole growth rate spectrum. To test whether fast growth was associated with diseases, the age–body mass relationship of these patients was tested, in a retrospective evaluation, for additional influence factors, such as dietary history and occurrence of certain diet and growth‐related diseases. No indication was found that animals particularly heavy for their age were more prone to diet/growth‐related disorders. In general, tortoises fed diets with meat/grain were heavier for their age than tortoises fed more appropriate diets; dietary history was not related to a particular disease. The results suggest the age–body mass relationship may not be suitable for testing effects of fast growth; an age–body length relationship would be more appropriate. Animals presented for a diet/growth‐related disorder were younger than animals presented for other reasons; there was a significant negative correlation between the severity of pyramiding and age, suggesting that growth‐related disorders may well limit the life expectancy of tortoises. Controlled clinical studies are required to fully test this hypothesis. Zoo Biol. 31:705‐717, 2012. © 2012 Wiley Periodicals, Inc.     

Histamine action on vertebrate GABAA receptors: direct channel gating and potentiation of GABA responses

Histamine is not only a crucial cytokine in the periphery but also an important neurotransmitter and neuromodulator in the brain. It is known to act on metabotropic H1-H4 receptors, but the existence of directly histamine-gated chloride channels in mammals has been suspected for many years. However, the molecular basis of such mammalian channels remained elusive, whereas in invertebrates, genes for histamine-gated channels have been already identified. In this report, we demonstrated that histamine can directly open vertebrate ion channels and identified beta subunits of GABA(A) receptors as potential candidates for histamine-gated channels. In Xenopus oocytes expressing homomultimeric beta channels, histamine evoked currents with an EC(50) of 212 microm (beta(2)) and 174 microm (beta(3)), whereas GABA is only a very weak partial agonist. We tested several known agonists and antagonists for the histamine-binding site of H1-H4 receptors and described for beta channels a unique pharmacological profile distinct from either of these receptors. In heteromultimeric channels composed of alpha(1)beta(2) or alpha(1)beta(2)gamma(2) subunits, we found that histamine is a modulator of the GABA response rather than an agonist as it potentiates GABA-evoked currents in a gamma(2) subunit-controlled manner. Despite the vast number of synthetic modulators of GABA(A) receptors widely used in medicine, which act on several distinct sites, only a few endogenous modulators have yet been identified. We show here for the first time that histamine modulates heteromultimeric GABA(A) receptors and may thus represent an endogenous ligand for an allosteric site.