Faculty Bibliography

Amyloid plaques are a hallmark of Alzheimer disease, but their importance in its pathogenesis is controversial. By neuronal labeling and transcranial two-photon imaging, we show in a transgenic mouse model of Alzheimer disease that dendrites passing through or near fibrillar amyloid deposits undergo spine loss and shaft atrophy, and nearby axons develop large varicosities, together leading to neurite breakage and large-scale, permanent disruption of neuronal connections. Thus, fibrillar amyloid deposition is more detrimental to neuronal circuitry than previously thought, underscoring the importance of prevention and early clearance of plaques

One of the major limitations in the current set of techniques available to neuroscientists is a dearth of methods for imaging individual cells deep within the brains of live animals. To overcome this limitation, we developed two forms of minimally invasive fluorescence microendoscopy and tested their abilities to image cells in vivo. Both one- and two-photon fluorescence microendoscopy are based on compound gradient refractive index (GRIN) lenses that are 350-1,000 microm in diameter and provide micron-scale resolution. One-photon microendoscopy allows full-frame images to be viewed by eye or with a camera, and is well suited to fast frame-rate imaging. Two-photon microendoscopy is a laser-scanning modality that provides optical sectioning deep within tissue. Using in vivo microendoscopy we acquired video-rate movies of thalamic and CA1 hippocampal red blood cell dynamics and still-frame images of CA1 neurons and dendrites in anesthetized rats and mice. Microendoscopy will help meet the growing demand for in vivo cellular imaging created by the rapid emergence of new synthetic and genetically encoded fluorophores that can be used to label specific brain areas or cell classes.

The GLP-1 receptor is a Class B heptahelical G-protein-coupled receptor that stimulates cAMP production in pancreatic beta-cells. GLP-1 utilizes this receptor to activate two distinct classes of cAMP-binding proteins: protein kinase A (PKA) and the Epac family of cAMP-regulated guanine nucleotide exchange factors (cAMPGEFs). Actions of GLP-1 mediated by PKA and Epac include the recruitment and priming of secretory granules, thereby increasing the number of granules available for Ca(2+)-dependent exocytosis. Simultaneously, GLP-1 promotes Ca(2+) influx and mobilizes an intracellular source of Ca(2+). GLP-1 sensitizes intracellular Ca(2+) release channels (ryanodine and IP (3) receptors) to stimulatory effects of Ca(2+), thereby promoting Ca(2+)-induced Ca(2+) release (CICR). In the model presented here, CICR activates mitochondrial dehydrogenases, thereby upregulating glucose-dependent production of ATP. The resultant increase in cytosolic [ATP]/[ADP] concentration ratio leads to closure of ATP-sensitive K(+) channels (K-ATP), membrane depolarization, and influx of Ca(2+) through voltage-dependent Ca(2+) channels (VDCCs). Ca(2+) influx stimulates exocytosis of secretory granules by promoting their fusion with the plasma membrane. Under conditions where Ca(2+) release channels are sensitized by GLP-1, Ca(2+) influx also stimulates CICR, generating an additional round of ATP production and K-ATP channel closure. In the absence of glucose, no 'fuel' is available to support ATP production, and GLP-1 fails to stimulate insulin secretion. This new 'feed-forward' hypothesis of beta-cell stimulus-secretion coupling may provide a mechanistic explanation as to how GLP-1 exerts a beneficial blood glucose-lowering effect in type 2 diabetic subjects

The development of spatial vision is relatively well documented in human and nonhuman primates. However, little is known about the development of sensitivity to motion. We measured the development of sensitivity to direction of motion, and the relationship between motion and contrast sensitivity in macaque monkeys as a function of age. Monkeys (Macaca nemestrina, aged between 10 days and 3 years) discriminated direction of motion in random-dot kinematograms. The youngest monkeys showed directionally selective orienting and the ability to integrate motion signals at large dot displacements and fast speeds. With age, coherence sensitivity improved for all spatial and temporal dot displacements tested. The temporal interval between the dots was far less important than the spatial offset in determining the animals' performance at all but the youngest ages. Motion sensitivity improved well beyond the end of the first postnatal year, when mid-spatial-frequency contrast sensitivity reached asymptote, and continued for at least 3 years. Sensitivity to contrast at high spatial frequencies also continued to develop beyond the end of the first year. We conclude that the development of motion sensitivity depends on mechanisms beyond the low-level filters presumed to limit acuity and contrast sensitivity, and most likely reflects the function of extrastriate visual areas

The rbcL sequences of 106 specimens representing 28 species of the four recognized sections of Orobanche were analyzed and compared. Most sequences represent pseudogenes with premature stop codons. This study confirms that the American lineage (sects. Gymnocaulis and Myzorrhiza) contains potentially functional rbcL-copies with intact open reading frames and low rates of non-synonymous substitutions. For the first time, this is also shown for a member of the Eurasian lineage, O. coerulescens of sect. Orobanche, while all other investigated species of sects. Orobanche and Trionychon contain pseudogenes with distorted reading frames and significantly higher rates of non-synonymous substitutions. Phylogenetic analyses of the rbcL sequences give equivocal results concerning the monophyly of Orobanche, and the American lineage might be more closely related to Boschniakia and Cistanche than to the other sections of Orobanche. Additionally, species of sect. Trionychon phylogenetically nest in sect. Orobanche. This is in concordance with results from other plastid markers (rps2 and matK), but in disagreement with other molecular (nuclear ITS), morphological, and karyological data. This might indicate that the ancestor of sect. Trionychon has captured the plastid genome, or parts of it, of a member of sect. Orobanche. Apart from the phylogenetically problematic position of sect. Trionychon, the phylogenetic relationships within sect. Orobanche are similar to those inferred from nuclear ITS data and are close to the traditional groupings traditionally recognized based on morphology. The intraspecific variation of rbcL is low and is neither correlated with intraspecific morphological variability nor with host range. Ancestral character reconstruction using parsimony suggests that the ancestor of O. sect. Orobanche had a narrow host range

Ultrasmall superparamagnetic iron oxide nanoparticles have been widely used during the past decade as MR intravascular contrast agents in the study of animal models. Such agents enhance both T1 and T2/T2* relaxation, although for animal studies it is the later type of enhancement that is most commonly exploited. Their strong microscopic intravascular susceptibility effect enables the local blood volume distribution to be mapped in various organs. High spatial resolution and sensitivity can be achieved, because the long half-life of these agents in blood, combined with anesthetization, permits steady-state measurements over extended periods. This capability has been utilized to study the cerebrovascular blood volume distributions and their changes in normal, activated, pathologic and pharmacologically or genetically modified states, particularly in rodent animal models. It has also been applied to study blood volume changes in other tissues, such as the myocardium. The relaxation rate shifts Delta R2 and Delta R2* induced by iron oxide agents may differ depending on certain morphological characteristics of the microvascular network, and sensitive Delta R2 and Delta R2* mapping can potentially provide, in addition to blood volume, measurement of other important microvascular parameters such as blood vessel density and size. This work aims to review the applications of ultrasmall superparamagnetic iron oxide contrast agents in MR animal studies, with an emphasis on the investigation of microvascular parameters

Magnetic resonance imaging (MRI) is a powerful method for in vivo quantification of tissue properties. It has been previously proposed that the index Q identical with Delta R2/(Delta R2*)2/3, where Delta R2 and Delta R2* are the spin echo and gradient echo relaxation rate shifts caused by the injection of an intravascular contrast agent, may be useful for characterizing microvasculature. In particular, Q is expected to correlate well with the density of microvessels. This study presents high-resolution in vivo Q-maps of normal mouse brain obtained with a superparamagnetic iron oxide contrast agent at a field of level of 9.4 T. Normative Q values are derived for several regions of interest and significant interregional variations are observed. Microvessel densities estimated from the Q-maps are found to be in reasonable accord with histologically determined values. A possible application of Q-maps is the assessment of angiogenic activity in tumors

The cerebellum consists of a highly organized set of folia that are largely generated postnatally during expansion of the granule cell precursor (GCP) pool. Since the secreted factor sonic hedgehog (Shh) is expressed in Purkinje cells and functions as a GCP mitogen in vitro, it is possible that Shh influences foliation during cerebellum development by regulating the position and/or size of lobes. We studied how Shh and its transcriptional mediators, the Gli proteins, regulate GCP proliferation in vivo, and tested whether they influence foliation. We demonstrate that Shh expression correlates spatially and temporally with foliation. Expression of the Shh target gene Gli1 is also highest in the anterior medial cerebellum, but is restricted to proliferating GCPs and Bergmann glia. By contrast, Gli2 is expressed uniformly in all cells in the developing cerebellum except Purkinje cells and Gli3 is broadly expressed along the anteroposterior axis. Whereas Gli mutants have a normal cerebellum, Gli2 mutants have greatly reduced foliation at birth and a decrease in GCPs. In a complementary study using transgenic mice, we show that overexpressing Shh in the normal domain does not grossly alter the basic foliation pattern, but does lead to prolonged proliferation of GCPs and an increase in the overall size of the cerebellum. Taken together, these studies demonstrate that positive Shh signaling through Gli2 is required to generate a sufficient number of GCPs for proper lobe growth