RESUMEN
Deep brain stimulation (DBS) is being investigated for a number of psychiatric indications, including posttraumatic stress disorder (PTSD). Preclinical studies continue to be a cornerstone for the development of new DBS applications. We investigate whether DBS delivered to the infralimbic cortex (IL), a region involved in mechanisms of stress resiliency, may counter behavioral abnormalities in rats that present persistent extinction deficits and long-term anxiety after exposure to fear conditioning. Rats undergoing fear conditioning/extinction were segregated into weak and strong extinction groups (WE >70% or SE <30% of freezing during extinction). Following 2 weeks of DBS, animals were exposed to novel recall sessions and tested in the open field, novelty-suppressed feeding, and elevated plus maze. zif268 expression was measured in structures involved in mechanisms of fear and stress. In vivo electrophysiology was used to record activity from the basolateral amygdala (BLA). We found that DBS improved extinction deficits and anxiety-like behavior in WE animals, having no significant effects in SE rats. No major differences in absolute zif268 levels were recorded across groups. However, correlation between zif268 expression in the IL and BLA was disrupted in WE animals, a deficit that was countered by DBS treatment. Electrophysiology experiments have shown that DBS reduced BLA firing of both putative principal cells and interneurons in WE rats, with no significant differences being detected between SE and SE DBS animals. In summary, IL DBS mitigated fear, partially improved anxiety-like behavior, reversed neurocircuitry abnormalities, and reduced BLA cell firing in a preclinical model of PTSD.
Asunto(s)
Ansiedad/fisiopatología , Complejo Nuclear Basolateral/fisiología , Estimulación Encefálica Profunda , Miedo/fisiología , Corteza Prefrontal/fisiología , Trastornos por Estrés Postraumático/fisiopatología , Animales , Complejo Nuclear Basolateral/metabolismo , Conducta Animal/fisiología , Condicionamiento Psicológico/fisiología , Proteína 1 de la Respuesta de Crecimiento Precoz/biosíntesis , Extinción Psicológica/fisiología , Pérdida de Tono Postural/fisiología , Masculino , Corteza Prefrontal/metabolismo , RatasRESUMEN
INTRODUCTION: Few, if any, radiotracers are available for the in vivo imaging of reactive oxygen species (ROS) in the central nervous system. ROS play a critical role in normal cell processes such as signaling and homeostasis but overproduction of ROS is implicated in several disorders. We describe here the radiosynthesis and initial ex vivo and in vivo evaluation of [11C]hydromethidine ([11C]HM) as a radiotracer to image ROS using positron emission tomography (PET). METHODS: [11C]HM and its deuterated isotopologue [11C](4) were produced using [11C]methyl triflate in a one-pot, two-step reaction and purified by high performance liquid chromatography. Ex vivo biodistribution studies were performed after tail vein injections of both radiotracers. To demonstrate sensitivity of uptake to ROS, [11C]HM was administered to rats treated systemically with lipopolysaccharide (LPS). In addition, ex vivo autoradiography and in vivo PET imaging were performed using [11C]HM on rats which had been microinjected with sodium nitroprusside (SNP) to induce ROS. RESULTS: [11C]HM and [11C](4) radiosyntheses were reliable and produced the radiotracers at high specific activities and radiochemical purities. Both radiotracers demonstrated good brain uptake and fast washout of radioactivity, but [11C](4) washout was faster. Pretreatment with LPS resulted in a significant increase in brain retention of radioactivity. Ex vivo autoradiography and PET imaging of rats unilaterally treated with microinjections of SNP demonstrated increased retention of radioactivity in the treated side of the brain. CONCLUSIONS: [11C]HM has the attributes of a radiotracer for PET imaging of ROS in the brain including good brain penetration and increased retention of radioactivity in animal models of oxidative stress.
Asunto(s)
Encéfalo/diagnóstico por imagen , Fenantridinas , Tomografía de Emisión de Positrones/métodos , Especies Reactivas de Oxígeno/metabolismo , Animales , Transporte Biológico , Encéfalo/metabolismo , Radioisótopos de Carbono , Fenantridinas/metabolismo , Fenantridinas/farmacocinética , Trazadores Radiactivos , Ratas , Distribución TisularRESUMEN
Assessments of total anticholinergic activity (SAA) in serum are of considerable interest for its potential involvement in cognitive impairment associated with polydrug states in the elderly and other populations. Such estimations have been based on the displacement of radioligand binding in rat brain tissues. The validity of such measurements has been questioned, as a potentially distorting effect of large serum proteins was identified. We sought to develop a modified assay that would be more efficient and free of this potential confound. Cultured CHO cells stably expressing M1 receptors M1WT3 were used. Binding of 3H-radioligands was conducted in 96-well plates and tested in serum containing known amounts of anticholinergic medications. Effects of endogenous serum proteins were assessed by pre-assay filtration and also by deproteinization with perchloric acid (PCA). Binding of [3H]quinuclidinyl benzilate ([3H]QNB) or [3H]N-methyl-scopolamine ([3H]NMS) to M1WT3 cells proved reliable and equally sensitive to varying concentrations of anticholinergic agents. In agreement with previous findings (Cox, Kwatra, Shetty, & Kwatra, 2009), filtration of proteins heavier than 50kDa essentially reduced SAA values to zero. In contrast, PCA preserved more than 70% of the binding seen untreated cell membranes. Cell-based assays also showed significant signal increases compared to the conventional rat brain-based protocol. Further advantages of the cell-based protocol described here include increased sensitivity and reliability, smaller amounts of radioligand needed, and higher throughput. PCA pretreatment eliminates potential artifacts attributable to serum proteins. This step, together with improvements in efficiency, should contribute significantly to the usefulness of the assay.