Mapping alterations to the endogenous elemental distribution within the lateral ventricles and choroid plexus in brain disorders using X-ray fluorescence imaging

The choroid plexus and cerebral ventricles are critical structures for the production of cerebral spinal fluid (CSF) and play an important role in regulating ion and metal transport in the brain, however many aspects of its roles in normal physiology and disease states, such as psychiatric illness, remain unknown. The choroid plexus is difficult to examine in vivo, and in situ ex vivo, and as such has typically been examined indirectly with radiolabeled tracers or ex vivo stains, making measurements of the endogenous K+, Cl-, and Ca+ distributions unreliable. In the present study, we directly examined the distribution of endogenous ions and biologically relevant transition metals in the choroid plexus and regions surrounding the ventricles (ventricle wall, cortex, corpus callosum, striatum) using X-ray fluorescence imaging (XFI). We find that the choroid plexus was rich in Cl- and Fe while K+ levels increase further from the ventricle as Cl- levels decrease, consistent with the known role of ion transporters in the choroid plexus CSF production. A polyI:C offspring displayed enlarged ventricles, elevated Cl- surrounding the ventricles, and intraventricular calcifications. These observations fit with clinical findings in patients with schizophrenia and suggest maternal treatment with polyI:C may lead to dysfunctional ion regulation in offspring. This study demonstrates the power of XFI for examining the endogenous elemental distributions of the ventricular system in healthy brain tissue as well as disease models

Routine cresyl violet (CV) histology and XFI elemental mapping (P, S, Cl, K, Ca, Fe, Cu, Zn) of the offspring (PND60) from a saline control rat and a polyI:C immune compromised rat.

<p>Images were collected from tissue sections -0.5 mm and -3.6 mm anterior to bregma. White arrows indicate the presence of numerous calcifications observed within the ventricles of the polyI;C offspring. Due to the increased swelling of the ventricle in the polyI:C offspring at bregma location -0.5mm, the medial side of the ventricle wall tore from the tissue section. A white dashed line shows the approximate location of the ventricle wall before it tore away during tissue sectioning. Scale bar = 500 μm, intensity units are μg cm^-2.</p

An enlarged view of elemental maps of Cl⁻, K⁺ and Cu in the choroid plexus and ventricle wall with tri-colour overlay.

<p>Numerous Cu hot spots mark the ventricle wall, and Cl⁻ and K⁺ co-localize with the choroid plexus epithelium. The tri-colour overlay highlights that K⁺ is at lower concentration outside the choroid plexus epithelium, while Cl⁻ is still abundant. Scale bar = 100 μm.</p

Cresyl violet (CV) histology and XFI elemental mapping of calcifications (white arrows) within ventricles.

<p>The first two columns correspond to images collected from tissue sections at a location of -0.5 mm anterior to bregma, and the last two columns correspond to images collected from tissue sections at a location of -3.6 mm anterior to bregma. Scale bar = 100 μm, intensity units are μg cm^-2.</p

Elemental quantification performed with XFI of distinct brain regions (choroid plexus, corpus callosum, striatum, cortex, and ventricle wall).

<p>One-way repeated measures ANOVA revealed significant differences in all elements as a factor of brain region (p<0.05). Tukey’s HSD was used to calculate post hoc tests. [<b>A</b>] Elemental concentrations for P, S, Cl, and K. A letter system is used to denote significant differences in P across brain regions. A = significantly different from choroid plexus, B = significantly different from corpus callosum, C = significantly different from striatum, D = significantly different from cortex, E = significantly different from ventricle wall. For all other elements, significant differences are indicated with an asterisk (*). [<b>B</b>] Elemental concentrations for Ca, Fe, Cu, and Zn. Significant differences are indicated with an asterisk (*).</p

Secondary Degeneration Impairs Myelin Ultrastructural Development in Adulthood following Adolescent Neurotrauma in the Rat Optic Nerve

Adolescence is a critical period of postnatal development characterized by social, emotional, and cognitive changes. These changes are increasingly understood to depend on white matter development. White matter is highly vulnerable to the effects of injury, including secondary degeneration in regions adjacent to the primary injury site which alters the myelin ultrastructure. However, the impact of such alterations on adolescent white matter maturation is yet to be investigated. To address this, female piebald-virol-glaxo rats underwent partial transection of the optic nerve during early adolescence (postnatal day (PND) 56) with tissue collection two weeks (PND 70) or three months later (PND 140). Axons and myelin in the transmission electron micrographs of tissue adjacent to the injury were classified and measured based on the appearance of the myelin laminae. Injury in adolescence impaired the myelin structure in adulthood, resulting in a lower percentage of axons with compact myelin and a higher percentage of axons with severe myelin decompaction. Myelin thickness did not increase as expected into adulthood after injury and the relationship between the axon diameter and myelin thickness in adulthood was altered. Notably, dysmyelination was not observed 2 weeks postinjury. In conclusion, injury in adolescence altered the developmental trajectory, resulting in impaired myelin maturation when assessed at the ultrastructural level in adulthood

Altered object exploration but not temporal order memory retrieval in an object recognition test following treatment of rats with the group II metabotropic glutamate receptor agonist LY379268

Temporal order memory refers to the ability to distinguish past experiences in the order that they occurred. Temporal order memory for objects is often tested in rodents using spontaneous object recognition paradigms. The circuitry mediating memory in these tests is distributed and involves ionotropic glutamate receptors in the perirhinal cortex and medial prefrontal cortex. It is unknown what role, if any, metabotropic glutamate receptors have in temporal order memory for objects. The present experiment examined the role of metabotropic glutamate receptors in temporal memory retrieval using the group II metabotropic glutamate receptor selective agonist LY379268. Rats were trained on a temporal memory test with three phases: two sample phases (60. min between them) in which rats explored two novel objects and a test phase (60. min after the second sample phase) which included a copy of each object previously encountered. Under these conditions, we confirmed that rats showed a significant exploratory preference for the object presented during the first sample phase. In a second experiment, we found that LY379268 (0.3, 1.0, or 3.0. mg/kg; i.p.; 30. min before the test phase) had no effect on temporal memory retrieval but dose-dependently reduced time spent exploring the objects. Our results show that enhancing mGluR2 activity under conditions when TM is intact does not influence memory retrieval. © 2013 Elsevier Ireland Ltd