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Purpose: To examine the important role that primary care nurse practitioners (NPs) have in providing long-term surveillance and health maintenance for breast, prostate, and colorectal cancer survivors throughout the continuum of cancer care.

Melatonin supplementation has been used as a therapeutic agent for several diseases, yet little is known about the underlying mechanisms by which melatonin synchronizes circadian rhythms. G-protein signaling plays a large role in melatonin-induced phase shifts of locomotor behavior and melatonin receptors activate G-protein-coupled inwardly rectifying potassium (GIRK) channels in Xenopus oocytes. The present study tested the hypothesis that melatonin influences circadian phase and electrical activity within the central clock in the suprachiasmatic nucleus (SCN) through GIRK channel activation. Unlike wild-type littermates, GIRK2 knock-out (KO) mice failed to phase advance wheel-running behavior in response to 3 d subcutaneous injections of melatonin in the late day. Moreover, in vitro phase resetting of the SCN circadian clock by melatonin was blocked by coadministration of a GIRK channel antagonist tertiapin-q (TPQ). Loose-patch electrophysiological recordings of SCN neurons revealed a significant reduction in the average action potential rate in response to melatonin. This effect was lost in SCN slices treated with TPQ and SCN slices from GIRK2 KO mice. The melatonin-induced suppression of firing rate corresponded with an increased inward current that was blocked by TPQ. Finally, application of ramelteon, a potent melatonin receptor agonist, significantly decreased firing rate and increased inward current within SCN neurons in a GIRKdependent manner. These results are the first to show that GIRK channels are necessary for the effects of melatonin and ramelteon within the SCN. This study suggests that GIRK channels may be an alternative therapeutic target for diseases with evidence of circadian disruption, including aberrant melatonin signaling.

The mammalian circadian and sleep-wake systems are closely aligned through their coordinated regulation of daily activity patterns. Although they differ in their anatomical organization and physiological processes, they utilize overlapping regulatory mechanisms that include an assortment of proteins and molecules interacting within the extracellular space. These extracellular factors include proteases that interact with soluble proteins, membrane-attached receptors and the extracellular matrix; and cell adhesion molecules that can form complex scaffolds connecting adjacent neurons, astrocytes and their respective intracellular cytoskeletal elements. Astrocytes also participate in the dynamic regulation of both systems through modulating neuronal appositions, the extracellular space and/or through release of gliotransmitters that can further contribute to the extracellular signaling processes. Together, these extracellular elements create a system that integrates rapid neurotransmitter signaling across longer time scales and thereby adjust neuronal signaling to reflect the daily fluctuations fundamental to both systems. Here we review what is known about these extracellular processes, focusing specifically on areas of overlap between the two systems. We also highlight questions that still need to be addressed. Although we know many of the extracellular players, far more research is needed to understand the mechanisms through which they modulate the circadian and sleep-wake systems.

Glutamate phase shifts the circadian clock in the mammalian suprachiasmatic nucleus (SCN) by activating NMDA receptors. Tissue-type plasminogen activator (tPA) gates phase shifts by activating plasmin to generate m(ature) BDNF, which binds TrkB receptors allowing clock phase shifts. Here, we investigate phase shifting in tPA knockout (tPA ; B6.129S2-Plat /J) mice, and identify urokinase-type plasminogen activator (uPA) as an additional circadian clock regulator. Behavioral activity rhythms in tPA mice entrain to a light-dark (LD) cycle and phase shift in response to nocturnal light pulses with no apparent loss in sensitivity. When the LD cycle is inverted, tPA mice take significantly longer to entrain than C57BL/6J wild-type (WT) mice. SCN brain slices from tPA mice exhibit entrained neuronal activity rhythms and phase shift in response to nocturnal glutamate with no change in dose-dependency. Pre-treating slices with the tPA/uPA inhibitor, plasminogen activator inhibitor-1 (PAI-1), inhibits glutamate-induced phase delays in tPA slices. Selective inhibition of uPA with UK122 prevents glutamate-induced phase resetting in tPA but not WT SCN slices. tPA expression is higher at night than the day in WT SCN, while uPA expression remains constant in WT and tPA slices. Casein-plasminogen zymography reveals that neither tPA nor uPA total proteolytic activity is under circadian control in WT or tPA SCN. Finally, tPA SCN tissue has lower mBDNF levels than WT tissue, while UK122 does not affect mBDNF levels in either strain. Together, these results suggest that either tPA or uPA can support photic/glutamatergic phase shifts of the SCN circadian clock, possibly acting through distinct mechanisms.

Background Cognitive decline in Alzheimer’s disease correlates with the spreading of neurofibrillary tangles consisting of aggregated forms of tau from the entorhinal cortex to synaptically connected brain regions. It is unclear how tau aggregation propagates across the brain but may involve trans‐synaptic transfer of tau between neurons or uptake of extracellular tau that is constitutively secreted from neurons. Specific receptors involved in these processes have remained elusive, and here we investigate the hypothesis that the LDL receptor‐related protein 1 (LRP1) functions as an endocytic receptor for tau. Method We employed 125I‐labeled tau to quantify its cellular uptake, confocal microscopy to examine its colocalization with LRP1, and surface plasmon resonance to assess direct binding of tau and phosphorylated forms of tau to LRP1. Result Our results reveal that monomeric tau binds LRP1 with high affinity, while curiously, phosphorylated forms of tau bind LRP1 with much lower affinity. The studies confirm that LRP1 internalization is attenuated the LRP1 antagonist RAP and is reduced by 70% in LRP1‐deficient cells. Confocal microscopy reveals that internalized tau colocalizes with LRP1 in endocytic vesicles. Most of the LRP1‐mediated uptake of tau leads to its lysosomal degradation. We also demonstrate that apolipoprotein E (ApoE) inhibits LRP1‐mediated tau internalization, with the ApoE4 isoform being a more potent inhibitor than other ApoE isoforms. Our studies in LRP1‐deficient cell lines confirm that a RAP‐insensitive second internalization mechanism occurs for tau, suggesting the existence of an LRP1‐independent receptor‐mediated process for tau uptake. Conclusion LRP1 functions as an endocytic receptor for tau, mediating the uptake of monomeric forms of tau resulting in its degradation. The ability of ApoE4 to modulate LRP1‐mediated uptake of tau may disrupt the normal physiological process of tau catabolism, leading to tau uptake by other mechanisms causing the spreading of neurofibrillary tangles and exacerbating the progression of tau‐mediated cognitive loss.

BackgroundThe spread of neurofibrillary tangle (NFT) pathology across the brain is a hallmark of Alzheimer’s disease (AD) and is thought to be driven by the cell‐to‐cell transport of seed competent tau protein. LRP1 is a receptor for tau that mediates tau uptake, degradation, and seeding of intracellular tau aggregation. We previously found that LRP1 binds recombinantly produced 2N4R tau with high affinity, and that phosphorylation on tau reduces its affinity for LRP1. However, it is unknown if LRP1 interacts with the soluble HMW forms of tau that drive tau seeding, and here we investigate LRP1 binding to HMW phospho‐mimetic tau or HMW tau isolated from AD patient brains.MethodWe recombinantly produced tau containing mutations to mimic phosphorylation patterns of high‐molecular weight (D20Q3) or low molecular weight (D8Q2) tau found in AD brains. Next, we measured their ability to bind purified LRP1 binding via surface plasmon resonance (SPR) and determined the ability of LRP1 expressed in cells to mediate 125I‐labeled tau internalization. We also investigate the binding of HMW pathogenic forms of tau as well as LMW forms of tau isolated by size exclusion chromatography from AD patient brain extracts to LRP1 using a novel SPR capture assay.ResultWe found that D20Q3 and D8Q2 tau bind LRP1 much weaker (Kd = 1249nM & Kd = 388nM, respectively) than unmodified 2N4R tau (Kd = 20nM). 125I‐labeled tau uptake assays indicate reduced uptake for both D20Q3 and D8Q2 tau compared to 2N4R tau. Finally, preliminary results demonstrate that LRP1 is able to bind pathogenic forms of HMW tau isolated from AD patient brains, although there appears to be variability from different patients and LRP1 may bind HMW tau weaker than LMW tau. Studies are underway to determine the affinity of these forms for LRP1.ConclusionOur results demonstrate that LRP1 binds pathogenic forms of tau, with significant variation seen between patients. Future studies will investigate if the affinity of these forms correlate with their ability to promote tau seeding.

In the mammalian circadian clock located in the suprachiasmatic nucleus (SCN) glutamate mimics light-induced phase shifts through activating NMDA receptors (NMDAR). These phase shifts require concurrent activation of TrkB receptors by brain-derived neurotrophic factor (BDNF). tPA contributes to phase shifting by influencing mBDNF generation. tPA also binds to receptors, including LRP-1, and may modulate phase shifting through non-proteolytic mechanisms related to such receptor binding. Here we investigate the effects of inhibiting LRP-1 on glutamate induced phase shifts, using either receptor associated protein (RAP) or an anti-LRP-1 polyclonal antibody to inhibit LRP-1 activity. SCN brain slices from adult male C57BL/6 mice were treated with glutamate (1mM) +/-RAP (500nM) or anti-LRP-1 antibody (75µg/mL) at ZT 16 or ZT 23 (where ZT 0=lights-on in the animal colony) for 10 minutes. The following day we recorded SCN single-unit neuronal activity (SUA) to determine the time of peak activity. Concurrent application of RAP or anti-LRP-1 with glutamate prevents the normal shifts, while RAP alone has no effect on clock phase, suggesting that in the SCN LRP-1 activity is important for glutamate-dependent clock phase regulation. We investigated the potential interaction between tPA and LRP-1 in SCN clock phase shifting using tPA knockout (KO) mice, B6.129S2-Plattm1Mlg/J. First we characterized the SUA rhythm in control tPA KO brain slices and after glutamate treatment. SCN slices from adult male tPA KO mice exhibit normally phased neuronal activity rhythms, and 10µM glutamate at ZT16 and ZT23 induces phase delays and phase advances, respectively. Thus, tPA KO mice do not exhibit severe deficiencies in clock phase regulation, possibly reflecting redundant mBDNF-generating pathways. Finally, RAP inhibits glutamate phase resetting in tPA KO slices, indicating LRP-1 does not act exclusively through interactions with tPA. Future studies will continue exploring tPA and LRP-1 regulation of SCN circadian clock phase.