Persistent cognitive deficits are one of the most important sequelae of head injury in humans. In an effort to model some of the structural and neuropharmacological changes that occur in chronic postinjury brains, we examined the longitudinal effects of moderate vertical controlled cortical impact (CCI) on place learning and memory using the Morris water maze (MWM) test, morphology, and vesicular acetylcholine (ACh) transporter (VAChT) and muscarinic receptor subtype 2 (M2) immunohistochemistry. Vertical CCI (left parietal cortex, 4 m/sec, 2.5 mm; n = 10) or craniotomy (sham) was produced in male Sprague-Dawley rats (n = 10). Place learning was tested at 2 weeks, 4 weeks, 3 months, 6 months, and 12 months postinjury with the escape platform in a different maze quadrant for each time point. At each interval, rats received 5 days of water maze acquisition (latency to find hidden platform), a probe trial to measure place memory, and 2 days of visible platform trials to control for nonspecific deficits. At 3 weeks, half the animals were sacrificed for histology. At these injury parameters, CCI produced no significant differences in place learning between injured and sham rats at 2 weeks, 4 weeks, or 6 months after injury. However, at 3 and 12 months, the injured rats took significantly longer to find the hidden platform than the sham rats. Probe trial performance differed only at 12 months postinjury between injured (25.73+/-2.1%, standard error of the mean) and sham rats (44.09+/-7.0%, p < 0.05). The maze deficits at 1 year were not due to a worsening of performance, but may have resulted from a reduced ability of injured rats to benefit from previous water maze experience. Hemispheric loss of 30.4+/-5.5 mm3 was seen at 3 weeks after injury (versus respective sham). However, hemispheric loss almost doubled by 1 year after injury (51.5+/-8.5 mm3, p < 0.05 versus all other groups). Progressive tissue loss was also reflected by a three- to fourfold increase in ipsilateral ventricular volume between 3 weeks and 1 year after injury. At 1 year after injury, immunostaining for VAChT was dramatically increased in all sectors of the hippocampus and cortex after injury. Muscarinic receptor subtype 2 (M2) immunoreactivity was dramatically decreased in the ipsilateral hippocampus. This suggests a compensatory response of cholinergic neurons to increase the efficiency of ACh neurotransmission. Moderate CCI in rats produces subtle MWM performance deficits accompanied by persistent alteration in M2 and VAChT immunohistochemistry and progressive tissue atrophy. The inability of injured rats to benefit from repeated exposures to the MWM may represent a deficit in procedural memory that is independent of changes in hippocampal cholinergic systems.

There is a broad discussion concerning the function of new neurons in the adult brain. An increasingly accepted hypothesis proposes their crucial role in spatial learning. In this work, however, we demonstrate adult cyclin D2 knockout (cD2 KO) mice, which lack adult hippocampal neurogenesis, are able to learn a spatial version of the Barnes maze. Similar to wild type (WT) controls, these mutant mice exhibited several indicators of learning during 6 days of training: successively shorter latency and distance, higher speed, and decreasing number of errors. WT and cD2 KO mice showed improved search strategies, which became increasingly spatial. During probe Trial 1, mutant mice attained the highest significant number of nose-pokes at the former target hole compared with all the other holes. Both WT and cD2 KO mice covered shorter distances during probe Trial 2, whereas the mutant mice showed higher speed. We also discuss the possibility that some of the observed differences displayed by cD2 KO mice during training and at the probe trials-for example, longer mean distance and more errors-are associated with a smaller hippocampal formation. Our results suggest that adult brain neurogenesis is not obligatory for learning the Barnes maze.

Stroke increases neurogenesis in the adult dentate gyrus in the short term, however, long-term effects at the cellular and functional level are poorly understood. Here we evaluated the impact of an early stroke lesion on neurogenesis and cognitive function of the aging brain. We hypothesized that a stroke disturbs dentate neurogenesis during aging correlate with impaired flexible learning. To address this issue a stroke was induced in 3-month-old C57Bl/6 mice by a middle cerebral artery occlusion (MCAO). To verify long-term changes of adult neurogenesis the thymidine analogue BrdU (5-Bromo-2′-deoxyuridine) was administrated at different time points during aging. One and half months after BrdU injections learning and memory performance were assessed with a modified version of the Morris water maze (MWM) that includes the re-learning paradigm, as well as hippocampus-dependent and -independent search strategies. After MWM performance mice were transcardially perfused. To further evaluate in detail the stroke-mediated changes on stem- and progenitor cells as well as endogenous proliferation nestin-green-fluorescent protein (GFP) mice were used. Adult nestin-GFP mice received a retroviral vector injection in the hippocampus to evaluate changes in the neuronal morphology. At an age of 20 month the nestin-GFP mice were transcardially perfused after MWM performance and BrdU application 1.5 months later. The early stroke lesion significantly decreased neurogenesis in 7.5- and 9-month-old animals and also endogenous proliferation in the latter group. Furthermore, immature doublecortin (DCX)-positive neurons were reduced in 20-month-old nestin-GFP mice after lesion. All MCAO groups showed an impaired performance in the MWM and mostly relied on hippocampal-independent search strategies. These findings indicate that an early ischemic insult leads to a dramatical decline of neurogenesis during aging that correlates with a premature development of hippocampal-dependent deficits. Our study supports the notion that an early stroke might lead to long-term cognitive deficits as observed in human patients after lesion.

Cortical spreading depolarizations are an epiphenomenon of human brain pathologies and associated with extensive but transient changes in ion homeostasis, metabolism, and blood flow. Previously, we have shown that cortical spreading depolarization have long-lasting consequences on the brains transcriptome and structure. In particular, we found that cortical spreading depolarization stimulate hippocampal cell proliferation resulting in a sustained increase in adult neurogenesis. Since the hippocampus is responsible for explicit memory and adult-born dentate granule neurons contribute to this function, cortical spreading depolarization might influence hippocampus-dependent cognition. To address this question, we induced cortical spreading depolarization in C57Bl/6 J mice by epidural application of 1.5 mol/L KCl and evaluated neurogenesis and behavior at two, four, or six weeks thereafter. Congruent with our previous findings in rats, we found that cortical spreading depolarization increases numbers of newborn dentate granule neurons. Moreover, exploratory behavior and object location memory were consistently enhanced. Reference memory in the water maze was virtually unaffected, whereas memory formation in the Barnes maze was impaired with a delay of two weeks and facilitated after four weeks. These data show that cortical spreading depolarization produces lasting changes in psychomotor behavior and complex, delay- and task-dependent changes in spatial memory, and suggest that cortical spreading depolarization-like events affect the emotional and cognitive outcomes of associated brain pathologies.

Aureochromes are light, oxygen, voltage (LOV) proteins and central blue-light receptors in algae acting as light-gated transcription factors. The C-terminal LOV domain mediates blue-light recognition and the basic region leucine zipper (bZIP) domain binds a specific DNA motif as effector. LOV domains from aureochromes have been successfully applied in optogenetic tools. The light-induced response of aureochromes has been studied by a variety of biophysical techniques, but the mechanism of signal progression from LOV to bZIP remains unclear. We studied the bZIP-LOV module of aureochrome1a from the diatom Phaeodactylum tricornutum using time-resolved rapid-scan FTIR difference spectroscopy. Time-resolved difference spectra of bZIP-LOV in vitro revealed a time constant of 5 s for the formation of a light state dimer of the LOV domains and the concomitant loss of α-helical elements in the bZIP domain. To verify these observations in a near-native environment, in-cell infrared difference spectroscopy (ICIRD) was extended from a steady state to a time-resolved technique using LOV domains in bacterial cells. We established a time-resolved in-cell method with a resolution of 7.6 ms after the laser pulse. Using this technique, the response of bZIP-LOV was followed in living bacterial cells and the light-induced partial unfolding of bZIP was confirmed to take place in cells in a similar time range as in vitro. These results provide structural and kinetic insights into the signaling mechanism of aureochromes. The slow response points to an association of LOV to bZIP in the dark state prior to activation.