The ApoE4 allele is associated with increased risk of small vessel disease, which is a cause of vascular cognitive impairment. Here, we report that mice with targeted replacement (TR) of the ApoE gene with human ApoE4 have reduced neocortical cerebral blood flow compared to ApoE3-TR mice, an effect due to reduced vascular density rather than slowing of microvascular red blood cell flow. Furthermore, homeostatic mechanisms matching local delivery of blood flow to brain activity are impaired in ApoE4-TR mice. In a model of cerebral hypoperfusion, these cerebrovascular alterations exacerbate damage to the white matter of the corpus callosum and worsen cognitive dysfunction. Using 3-photon microscopy we found that the increased white matter damage is linked to an enhanced reduction of microvascular flow resulting in local hypoxia. Such alterations may be responsible for the increased susceptibility to hypoxic-ischemic lesions in the subcortical white matter of individuals carrying the ApoE4 allele.

Phosphatidylinositol 3-kinase (PI3K) has been implicated in synaptic plasticity and other neural functions in the brain. However, the role of individual PI3K isoforms in the brain is unclear. We investigated the role of PI3Kγ in hippocampal-dependent synaptic plasticity and cognitive functions. We found that PI3Kγ has a crucial and specific role in NMDA receptor (NMDAR)-mediated synaptic plasticity at mouse Schaffer collateral-commissural synapses. Both genetic deletion and pharmacological inhibition of PI3Kγ disrupted NMDAR long-term depression (LTD) while leaving other forms of synaptic plasticity intact. Accompanying this physiological deficit, the impairment of NMDAR LTD by PI3Kγ blockade was specifically correlated with deficits in behavioral flexibility. These findings suggest that a specific PI3K isoform, PI3Kγ, is critical for NMDAR LTD and some forms of cognitive function. Thus, individual isoforms of PI3Ks may have distinct roles in different types of synaptic plasticity and may therefore influence various kinds of behavior.

No abstract

Hypertension is a leading cause of stroke and dementia, effects attributed to disrupting delivery of blood flow to the brain. Hypertension also alters the blood-brain barrier (BBB), a critical component of brain health. Although endothelial cells are ultimately responsible for the BBB, the development and maintenance of the barrier properties depend on the interaction with other vascular-associated cells. However, it remains unclear if BBB disruption in hypertension requires cooperative interaction with other cells. Perivascular macrophages (PVM), innate immune cells closely associated with cerebral microvessels, have emerged as major contributors to neurovascular dysfunction. Using 2-photon microscopy in vivo and electron microscopy in a mouse model of Ang II (angiotensin II) hypertension, we found that the vascular segments most susceptible to increased BBB permeability are arterioles and venules >10 µm and not capillaries. Brain macrophage depletion with clodronate attenuates, but does not abolish, the increased BBB permeability in these arterioles where PVM are located. Deletion of AT1R (Ang II type-1 receptors) in PVM using bone marrow chimeras partially attenuated the BBB dysfunction through the free radical-producing enzyme Nox2. In contrast, downregulation of AT1R in cerebral endothelial cells using a viral gene transfer-based approach prevented the BBB disruption completely. The results indicate that while endothelial AT1R, mainly in arterioles and venules, initiate the BBB disruption in hypertension, PVM are required for the full expression of the dysfunction. The findings unveil a previously unappreciated contribution of resident brain macrophages to increased BBB permeability of hypertension and identify PVM as a putative therapeutic target in diseases associated with BBB dysfunction.

Background and Purpose Cerebral microbleeds are linked to cognitive decline, but it remains unclear how they impair neuronal function. Infarction is not typically observed near microbleeds, suggesting more subtle mechanisms, such as inflammation, may play a role. Due to their small size and largely asymptomatic nature, real-time detection and study of spontaneous cerebral microbleeds in humans and animal models is difficult. Methods We used in vivo two-photon microscopy through a chronic cranial window in adult mice to follow the inflammatory response after a cortical microhemorrhage of ∼100-μm diameter, induced by rupturing a targeted cortical arteriole with a laser. Results The inflammatory response included the invasion of blood borne leukocytes, the migration and proliferation of brain resident microglia, and the activation of astrocytes. Nearly all inflammatory cells responding to the microhemorrhage were brain-resident microglia, but a small number of CX3CR1+ and CCR2+ macrophages, ultimately originating from the invasion of blood-borne monocytes, were also found near the lesion. We found a coordinated pattern of microglia migration and proliferation, where microglia within 200 μm of the microhemorrhage migrated toward the lesion over hours to days. In contrast, microglia proliferation was not observed until ∼40 hours after the lesion and occurred primarily in a shell-shaped region where the migration of microglia decreased their local density. This data suggests that local microglia density changes may trigger proliferation. Astrocytes activated in a similar region as microglia, but delayed by a few days. By two weeks, this inflammatory response had largely resolved. Conclusions While microhemorrhages are small in size, the brain responds to a single bleed with an inflammatory response that involves brain-resident and blood-derived cells, persists for weeks, and may impact the adjacent brain microenvironment.

The amyloid-b (Ab) peptide, a key pathogenic factor in Alzheimer's disease, attenuates the increase in cerebral blood flow (CBF) evoked by neural activity (functional hyperemia), a vital homeostatic response in which NMDA receptors (NMDARs) play a role through nitric oxide, and the CBF increase produced by endothelial factors. Tissue plasminogen activator (tPA), which is reduced in Alzheimer's disease and in mouse models of Ab accumulation, is required for the full expression of the NMDAR-dependent component of functional hyperemia. Therefore, we investigated whether tPA is involved in the neurovascular dysfunction of Ab. tPA activity was reduced, and the tPA inhibitor plasminogen inhibitor-1 (PAI-1) was increased in male mice expressing the Swedish mutation of the amyloid precursor protein (tg2576). Counteracting the tPA reduction with exogenous tPA or with pharmacological inhibition or genetic deletion of PAI-1 completely reversed the attenuation of the CBF increase evoked by whisker stimulation but did not ameliorate the response to the endothelium-dependent vasodilator acetylcholine. The tPA deficit attenuated functional hyperemia by suppressing NMDAR-dependent nitric oxide production during neural activity. Pharmacological inhibition of PAI-1 increased tPA activity, prevented neurovascular uncoupling, and ameliorated cognition in 11-to 12-month-old tg2576 mice, effects associated with a reduction of cerebral amyloid angiopathy but not amyloid plaques. The data unveil a selective role of the tPA in the suppression of functional hyperemia induced by Ab and in the mechanisms of cerebral amyloid angiopathy, and support the possibility that modulation of the PAI-1-tPA pathway may be beneficial in diseases associated with amyloid accumulation. Significance StatementAmyloid-b (Ab ) peptides have profound neurovascular effects that may contribute to cognitive impairment in Alzheimer's disease. We found that Ab attenuates the increases in blood flow evoked by neural activation through a reduction in tissue plasminogen activator (tPA) caused by upregulation of its endogenous inhibitor plasminogen inhibitor-1 (PAI-1). tPA deficiency prevents NMDA receptors from triggering nitric oxide production, thereby attenuating the flow increase evoked by neural activity. PAI-1 inhibition restores tPA activity, rescues neurovascular coupling, reduces amyloid deposition around blood vessels, and improves cognition in a mouse model of Ab accumulation. The findings demonstrate a previously unappreciated role of tPA in Ab -related neurovascular dysfunction and in vascular amyloid deposition. Restoration of tPA activity could be of therapeutic value in diseases associated with amyloid accumulation.

In the cerebellum, synaptic strength at the synapses between parallel fibers and Purkinje cells is best known to be modulated via metabotropic glutamate receptor 1 (mGluR1)-dependent cerebellar long-term depression (LTD). An increase in intracellular calcium levels plays an important role in inducing mGluR1-dependent cerebellar LTD. Downstream of mGluR1, there are two major sources of calcium: transient receptor potential canonical (TRPC) channels and inositol trisphosphate receptors (IP 3 R). IP 3 R triggers a calcium release from the intracellular calcium store. Here, we show that TRPC channels mediate mGluR1-evoked slow currents to regulate cerebellar LTD in Sprague Dawley rats. We found that the inhibition of TRPC channels blocks the induction of cerebellar LTD. Moreover, we show that processes known to underlie cerebellar LTD induction, such as increases in intracellular calcium concentration, the activation of protein kinase C, and the internalization of GluR2, are also hindered by blocking TRPC. These results suggest that the mGluR1-evoked activation of TRPC channels is required for the induction of cerebellar LTD.

We show that third harmonic generation (THG) microscopy using a 1-MHz train of 1,300-nm femtosecond duration laser pulses enabled visualization of the structure and quantification of flow speed in the cortical microvascular network of mice to a depth of > 1 mm. Simultaneous three-photon imaging of an intravascular fluorescent tracer enabled us to quantify the cell free layer thickness. Using the label-free imaging capability of THG, we measured flow speed in different types of vessels with and without the presence of an intravascular tracer conjugated to a high molecular weight dextran (2 MDa FITC-dextran, 5% w/v in saline, 100 µl). We found a ∼20% decrease in flow speeds in arterioles and venules due to the dextran-conjugated FITC, which we confirmed with Doppler optical coherence tomography. Capillary flow speeds did not change, although we saw a ∼7% decrease in red blood cell flux with dextran-conjugated FITC injection.