Optogenetic Manipulation of Cerebellar Purkinje Cell Activity In Vivo

Purkinje cells (PCs) are the sole output neurons of the cerebellar cortex. Although their anatomical connections and physiological response properties have been extensively studied, the causal role of their activity in behavioral, cognitive and autonomic functions is still unclear because PC activity cannot be selectively controlled. Here we developed a novel technique using optogenetics for selective and rapidly reversible manipulation of PC activity in vivo. We injected into rat cerebellar cortex lentiviruses expressing either the light-activated cationic channel channelrhodopsin-2 (ChR2) or light-driven chloride pump halorhodopsin (eNpHR) under the control of the PC-specific L7 promoter. Transgene expression was observed in most PCs (ChR2, 92.6%; eNpHR, 95.3%), as determined by immunohistochemical analysis. In vivo electrophysiological recordings showed that all light-responsive PCs in ChR2-transduced rats increased frequency of simple spike in response to blue laser illumination. Similarly, most light-responsive PCs (93.8%) in eNpHR-transduced rats decreased frequency of simple spike in response to orange laser illumination. We then applied these techniques to characterize the roles of rat cerebellar uvula, one of the cardiovascular regulatory regions in the cerebellum, in resting blood pressure (BP) regulation in anesthetized rats. ChR2-mediated photostimulation and eNpHR-mediated photoinhibition of the uvula had opposite effects on resting BP, inducing depressor and pressor responses, respectively. In contrast, manipulation of PC activity within the neighboring lobule VIII had no effect on BP. Blue and orange laser illumination onto PBS-injected lobule IX didn't affect BP, indicating the observed effects on BP were actually due to PC activation and inhibition. These results clearly demonstrate that the optogenetic method we developed here will provide a powerful way to elucidate a causal relationship between local PC activity and functions of the cerebellum

Bidirectional regulation of neurite elaboration by alternatively spliced metabotropic glutamate receptor 5 (mGlur5) isoforms

Alternative splicing in the mGluR5 gene generates two different receptor isoforms, of which expression is developmentally regulated. However, little is known about the functional significance of mGluR5 splice variants. We have examined the functional coupling, subcellular targeting, and effect on neuronal differentiation of epitope-tagged mGluR5 isoforms by expression in neuroblastoma NG108-15 cells. We found that both mGluR5 splice variants give rise to comparable [Ca2+]i transients and have similar pharmacological profile. Tagged receptors were shown by immunofluorescence to be inserted in the plasma membrane. In undifferentiated cells the subcellular localization of the two mGluR5 isoforms was partially segregated, whereas in differentiated cells the labeling largely redistributed to the newly formed neurites. Interestingly, we demonstrate that mGluR5 splice variants dramatically influence the formation and maturation of neurites; mGluR5a hinders the acquisition of mature neuronal traits and mGluR5b fosters the elaboration and extension of neurites. These effects are partly inhibited by MPEP

Paraesophageal hernia repair: a curative consideration for chronic anemia?

Introduction Chronic anemia is a common, coinciding or presenting diagnosis in patients with paraesophageal hernia (PEH). Presence of endoscopically identified ulcerations frequently prompts surgical consultation in the otherwise asymptomatic patient with anemia. Rates of anemia resolution following paraesophageal hernia repair (PEHR) often exceed the prevalence of such lesions in the study population. A defined algorithm remains elusive. This study aims to characterize resolution of anemia after PEHR with respect to endoscopic diagnosis. Materials and methods Retrospective review of a prospectively maintained database of patients with PEH and anemia undergoing PEHR from 2007 to 2018 was performed. Anemia was determined by preoperative labs: Hgb < 12 mg/dl in females, Hgb < 13 mg/dl in males, or patients with ongoing iron supplementation. Improvement of post-operative anemia was assessed by post-operative hemoglobin values and continued necessity of iron supplementation. Results Among 56 identified patients, 45 were female (80.4%). Forty patients (71.4%) were anemic by hemoglobin value, 16 patients (28.6%) required iron supplementation. Mean age was 65.1 years, with mean BMI of 27.7 kg/m 2 . One case was a Type IV PEH and the rest Type III. 32 (64.0%) had potential source of anemia: 16 (32.0%) Cameron lesions, 6 (12.0%) gastric ulcers, 12 (24.0%) gastritis. 10 (20.0%) had esophagitis and 4 (8%) Barrett’s esophagus. 18 (36%) PEH patients had normal preoperative EGD. Median follow-up was 160 days. Anemia resolution occurred in 46.4% of patients. Of the 16 patients with pre-procedure Cameron lesions, 10 (63%) had resolution of anemia. Patients with esophagitis did not achieve resolution. 72.2% (13/18) of patients with no lesions on EGD had anemia resolution ( p  = 0.03). Conclusion Patients with PEH and identifiable ulcerations showed 50% resolution of anemia after hernia repair. Patients without identifiable lesions on endoscopy demonstrated statistically significant resolution of anemia in 72.2% of cases. Anemia associated with PEH adds an indication for surgical repair with curative intent