Regulation of glucose homeostasis and insulin action by ceramide acyl-chain length: A beneficial role for very long-chain sphingolipid species.

In a recent study, we showed that in response to high fat feeding C57BL/6, 129X1, DBA/2 and FVB/N mice all developed glucose intolerance, while BALB/c mice displayed minimal deterioration in glucose tolerance and insulin action. Lipidomic analysis of livers across these five strains has revealed marked strain-specific differences in ceramide (Cer) and sphingomyelin (SM) species with high-fat feeding; with increases in C16-C22 (long-chain) and reductions in C>22 (very long-chain) Cer and SM species observed in the four strains that developed HFD-induced glucose intolerance. Intriguingly, the opposite pattern was observed in sphingolipid species in BALB/c mice. These strain-specific changes in sphingolipid acylation closely correlated with ceramide synthase 2 (CerS2) protein content and activity, with reduced CerS2 levels/activity observed in glucose intolerant strains and increased content in BALB/c mice. Overexpression of CerS2 in primary mouse hepatocytes induced a specific elevation in very long-chain Cer, but despite the overall increase in ceramide abundance, there was a substantial improvement in insulin signal transduction, as well as decreased ER stress and gluconeogenic markers. Overall our findings suggest that very long-chain sphingolipid species exhibit a protective role against the development of glucose intolerance and hepatic insulin resistance.

Activation of BK and SK channels by efferent synapses on outer hair cells in high-frequency regions of the rodent cochlea.

Cholinergic neurons of the brainstem olivary complex project to and inhibit outer hair cells (OHCs), refining acoustic sensitivity of the mammalian cochlea. In all vertebrate hair cells studied to date, cholinergic inhibition results from the combined action of ionotropic acetylcholine receptors and associated calcium-activated potassium channels. Although inhibition was thought to involve exclusively small conductance (SK potassium channels), recent findings have shown that BK channels also contribute to inhibition in basal, high-frequency OHCs after the onset of hearing. Here we show that the waveform of randomly timed IPSCs (evoked by high extracellular potassium) in high-frequency OHCs is altered by blockade of either SK or BK channels, with BK channels supporting faster synaptic waveforms and SK channels supporting slower synaptic waveforms. Consistent with these findings, IPSCs recorded from high-frequency OHCs that express BK channels are briefer than IPSCs recorded from low-frequency (apical) OHCs that do not express BK channels and from immature high-frequency OHCs before the developmental onset of BK channel expression. Likewise, OHCs of BKα(-/-) mice lacking the pore-forming α-subunit of BK channels have longer IPSCs than do the OHCs of BKα(+/+) littermates. Furthermore, serial reconstruction of electron micrographs showed that postsynaptic cisterns of BKα(-/-) OHCs were smaller than those of BKα(+/+) OHCs, and immunofluorescent quantification showed that efferent presynaptic terminals of BKα(-/-) OHCs were smaller than those of BKα(+/+) OHCs. Together, these findings indicate that BK channels contribute to postsynaptic function, and influence the structural maturation of efferent-OHC synapses.

Deletion of Shank1 has minimal effects on the molecular composition and function of glutamatergic afferent postsynapses in the mouse inner ear.

Shank proteins (1-3) are considered the master organizers of glutamatergic postsynaptic densities in the central nervous system, and the genetic deletion of either Shank1, 2, or 3 results in altered composition, form, and strength of glutamatergic postsynapses. To investigate the contribution of Shank proteins to glutamatergic afferent synapses of the inner ear and especially cochlea, we used immunofluorescence and quantitative real time PCR to determine the expression of Shank1, 2, and 3 in the cochlea. Because we found evidence for expression of Shank1 but not 2 and 3, we investigated the morphology, composition, and function of afferent postsynaptic densities from defined tonotopic regions in the cochlea of Shank1(-/-) mice. Using immunofluorescence, we identified subtle changes in the morphology and composition (but not number and localization) of cochlear afferent postsynaptic densities at the lower frequency region (8 kHz) in Shank1(-/-) mice compared to Shank1(+/+) littermates. However, we detected no differences in auditory brainstem responses at matching or higher frequencies. We also identified Shank1 in the vestibular afferent postsynaptic densities, but detected no differences in vestibular sensory evoked potentials in Shank1(-/-) mice compared to Shank1(+/+) littermates. This work suggests that Shank proteins play a different role in the development and maintenance of glutamatergic afferent synapses in the inner ear compared to the central nervous system.

Ring bands in fish skeletal muscle: reorienting the myofibrils and microtubule cytoskeleton within a single cell.

Skeletal muscle cells (fibers) contract by shortening their parallel subunits, the myofibrils. Here we show a novel pattern of myofibril orientation in white muscle fibers of large black sea bass, Centropristis striata. Up to 48% of the white fibers in fish >1168 g had peripheral myofibrils undergoing an ∼90(o) shift in orientation. The resultant ring band wrapped the middle of the muscle fibers and was easily detected with polarized light microscopy. Transmission electron microscopy showed that the reoriented myofibrils shared the cytoplasm with the central longitudinal myofibrils. A microtubule network seen throughout the fibers surrounded nuclei but was mostly parallel to the long-axis of the myofibrils. In the ring band portion of the fibers the microtubule cytoskeleton also shifted orientation. Sarcolemmal staining with anti-synapsin was the same in fibers with or without ring bands, suggesting that fibers with ring bands have normal innervation and contractile function. The ring bands appear to be related to body-mass or age, not fiber size, and also vary along the body, being more frequent at the midpoint of the anteroposterior axis. Similar structures have been reported in different taxa and appear to be associated with hypercontraction of fibers not attached to a rigid structure (bone) or with fibers with unusually weak links between the sarcolemma and cytoskeleton, as in muscular dystrophy. Fish muscle fibers are attached to myosepta, which are flexible and may allow for fibers to hypercontract and thus form ring bands. The consequences of such a ring band pattern might be to restrict the further expansion of the sarcolemma and protect it from further mechanical stress.