A single sub-anesthetic dose of ketamine exerts rapid and sustained antidepressant effects. Here, we examined the role of the ventral hippocampus (vHipp)-medial prefrontal cortex (mPFC) pathway in ketamine's antidepressant response. Inactivation of the vHipp with lidocaine prevented the sustained, but not acute, antidepressant-like effect of ketamine as measured by the forced swim test (FST). Moreover, optogenetic as well as pharmacogenetic specific activation of the vHipp-mPFC pathway using DREADDs (designer receptors exclusively activated by designer drugs) mimicked the antidepressant-like response to ketamine; importantly, this was pathway specific, in that activation of a vHipp to nucleus accumbens circuit did not do this. Furthermore, optogenetic inactivation of the vHipp/mPFC pathway at the time of FST completely reversed ketamine's antidepressant response. In addition, we found that a transient increase in TrkB receptor phosphorylation in the vHipp contributes to ketamine's sustained antidepressant response. These data demonstrate that activity in the vHipp-mPFC pathway is both necessary and sufficient for the antidepressant-like effect of ketamine.

Chronic intermittent hypoxia (CIH) models repetitive bouts of arterial hypoxemia that occur in humans suffering from obstructive sleep apnea. CIH has been linked to persistent activation of arterial chemoreceptors and the renin-angiotensin system, which have been linked to chronic elevations of sympathetic nerve activity (SNA) and mean arterial pressure (MAP). Because Fos and FosB are transcription factors involved in activator protein (AP)-1 driven central nervous system neuronal adaptations, this study determined if CIH causes increased Fos or FosB staining in brain regions that regulate SNA and autonomic function. Male Sprague Dawley rats were instrumented with telemetry transmitters for continuous recording of MAP and heart rate (HR). Rats were exposed to continuous normoxia (CON) or to CIH for 8 h/day for 7 days. CIH increased MAP by 7-10 mmHg without persistently affecting HR. A separate group of rats was killed 1 day after 7 days of CIH for immunohistochemistry. CIH did not increase Fos staining in any brain region examined. Staining for FosB/ΔFosB was increased in the organum vasculosum of the lamina terminalis (CON: 9 ± 1; CIH: 34 ± 3 cells/section), subfornical organ (CON: 7 ± 2; CIH: 31 ± 3), median preoptic nucleus (CON 15 ± 1; CIH: 38 ± 3), nucleus of the solitary tract (CON: 9 ± 2; CIH: 28 ± 4), A5 (CON: 3 ± 1; CIH: 10 ± 1), and rostral ventrolateral medulla (CON: 5 ± 1; CIH: 17 ± 2). In the paraventricular nucleus, FosB/ΔFosB staining was located mainly in the dorsal and medial parvocellular subnuclei. CIH did not increase FosB/ΔFosB staining in caudal ventrolateral medulla or supraoptic nucleus. These data indicate that CIH induces an increase in FosB/ΔFosB in autonomic nuclei and suggest that AP-1 transcriptional regulation may contribute to stable adaptive changes that support chronically elevated SNA.

Major depressive disorder (MDD) is prevalent. Although standards antidepressants are more effective than placebo, up to 35% of patients do not respond to 4 or more conventional treatments and are considered to have treatment-resistant depression (TRD). Considerable effort has been devoted to trying to find effective treatments for TRD. This review focuses on vagus nerve stimulation (VNS), approved for TRD in 2005 by the Food and Drugs Administration. Stimulation is carried by bipolar electrodes on the left cervical vagus nerve, which are attached to an implanted stimulator generator. The vagus bundle contains about 80% of afferent fibers terminating in the medulla, from which there are projections to many areas of brain, including the limbic forebrain. Various types of brain imaging studies reveal widespread functional effects in brain after either acute or chronic VNS. Although more randomized control trials of VNS need to be carried out before a definitive conclusion can be reached about its efficacy, the results of open studies, carried out over period of 1 to 2 years, show much more efficacy when compared with results from treatment as usual studies. There is an increase in clinical response to VNS between 3 and 12 months, which is quite different from that seen with standard antidepressant treatment of MDD. Preclinically, VNS affects many of the same brain areas, neurotransmitters (serotonin, norepinephrine) and signal transduction mechanisms (brain-derived neurotrophic factor-tropomyosin receptor kinase B) as those found with traditional antidepressants. Nevertheless, the mechanisms by which VNS benefits patients nonresponsive to conventional antidepressants is unclear, with further research needed to clarify this.

Mice harboring 1, 2, or 3 copies of the angiotensin-converting enzyme (ACE) gene were used to evaluate the quantitative role of the ACE locus on obesity. Three-copy mice fed with a high-fat diet had lower body weight and peri-epididymal adipose tissue than did 1- and 2-copy mice (P < 0.05). On regular diet, 3-copy mice had to eat more to maintain the same body weight; on a high-fat diet, they ate the same but weighed less than 1- and 2-copy mice (P < 0.05), indicating a higher metabolic rate in 3-copy mice that was not affected by ANG II AT(1) blocker treatment. A catalytically inactive form of thimet oligopeptidase (EC 3.4.24.15; EP24.15) was used to isolate ACE substrates from adipose tissue. Liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) identified 162 peptide peaks; 16 peptides were present in both groups (1- and 3-copy mice fed with a high-fat diet), whereas 58 of the 72 unique peptides were found only in the 3-copy mice. Peptide size distribution was shifted to lower molecular weight in 3-copy mice. Two of the identified peptides, LVVYPWTQRY and VVYPWTQRY, which are ACE substrates, inhibited in vitro protein kinase C phosphorylation in a concentration-dependent manner. In addition, neurolysin (EC 3.4.24.16; EP24.16) activity was lower in fat tissue from 3- vs. 1-copy mice (P < 0.05). Taken together, these results provide evidence that ACE is associated with body weight and peri-epididymal fat accumulation. This response may involve the generation of oligopeptides that inhibit the activity of EP24.16 and other oligopeptidases within the adipose tissue.

Inappropriate vasopressin (AVP) release causes dilutional hyponatremia in many pathophysiological states such as cirrhosis. The central molecular mechanisms that mediate inappropriate AVP release are unknown. We tested the hypothesis that changes in the expression or trafficking of TRPV4 in the central nervous system may contribute to inappropriate AVP release in the bile duct ligation (BDL) model of cirrhosis in the rat. Four weeks after surgery, BDL rats demonstrated significantly increased plasma vasopressin and plasma renin activity (PRA), hypervolemia, and decreased plasma osmolality. These effects were blocked by providing BDL rats with 2% saline to drink for 15 days. TRPV4 protein expression was significantly increased in brain punches from BDL rats containing the supraoptic nucleus (SON) of the hypothalamus (100% +/- 11 to 157% +/- 4.8), and this effect was blocked in BDL rats given saline. Immunohistochemistry demonstrated a significant increase in TRPV4-positive cells and the percentage of AVP neurons that also were TRPV4-positive in the SON of BDL rats. In the hypothalamus of BDL rats, TRPV4 lipid raft association increased compared with sham (from 100% +/- 2.1 to 326.1% +/- 16). This effect was significantly attenuated in BDL rats given 2% saline to drink (174% +/- 11). In the brain stem, TRPV4 lipid raft association was reduced by BDL and inversely related to plasma AVP and PRA. We speculate that changes in TRPV4 expression and compartmentalization within lipid rafts could contribute to a feed-forward mechanism related to AVP release in cirrhosis.

Endopeptidase 24.15 (ep24.15: EC3.4.24.15), a secreted protein involved in peptide metabolism, is unusual in that it does not contain a signal peptide sequence. In this work, we describe the physical interaction between ep24.15 and 14-3-3 epsilon, one isoform of a family of ubiquitous phosphoserine/threoninescaffold proteins that organizes cell signaling and is involved in exocytosis. The interaction between ep24.15 and 14-3-3 epsilon increased following phosphorylation of ep24.15 at Ser 644 by protein kinase A (PKA). The co-localization of ep24.15 and 14-3-3 epsilon was increased by exposure of HEK293 cells (human embryonic kidney cells) to forskolin (10 lM). Overexpression of 14-3-3 epsilon in HEK293 cells almost doubled the secretion of ep24.15 stimulated by A23187 (7.5 lM) from 10% [1.4 ± 0.24 AFU/(min 10 6 cells)] to 19% [2.54 ± 0.24 AFU/(min 10 6 cells)] (p < 0.001) of the total intracellular enzyme activity. Treatment with forskolin had a synergistic effect on the A23187-stimulated secretion of ep24.15 that was totally blocked by the PKA inhibitor KT5720. The ep24.15 point mutation S644A reduced the co-localization of ep24.15 and 14-3-3 in stably transfected HEK293 cells. Indeed, secretion of the ep24.15 S644A mutant from these cells was only slightly stimulated by A23187 and insensitive to forskolin, in contrast to that of the wild type enzyme. Together, these data suggest that prior interaction with 14-3-3 is an important step in the unconventional stimulated secretion of ep24.15.

BackgroundVagal nerve stimulation (VNS) has been approved for treatment-resistant depression. Many antidepressants increase expression of brain-derived neurotrophic factor (BDNF) in brain or activate, via phosphorylation, its receptor, TrkB. There have been no studies yet of whether VNS would also cause phosphorylation of TrkB.MethodsWestern blot analysis was used to evaluate the phosphorylation status of TrkB in the hippocampus of rats administered VNS either acutely or chronically. Acute effects of VNS were compared with those caused by fluoxetine or desipramine (DMI) whereas its chronic effects were compared with those of sertraline or DMI.ResultsAll treatments, given either acutely or chronically, significantly elevated phosphorylation of tyrosines 705 and 816 on TrkB in the hippocampus. However, only VNS increased the phosphorylation of tyrosine 515, with both acute and chronic administration causing this effect. Pretreatment with K252a, a nonspecific tyrosine kinase inhibitor, blocked the phosphorylation caused by acute VNS at all three tyrosines. Downstream effectors of Y515, namely Akt and ERK, were also phosphorylated after acute treatment with VNS, whereas DMI did not cause this effect.ConclusionVNS rapidly activates TrkB phosphorylation and this effect persists over time. VNS-induced phosphorylation of tyrosine 515 is distinct from the effect of standard antidepressant drugs.

We studied the effects of water deprivation on the phosphorylation of TrkB and NMDA receptor subunits in the supraoptic nucleus (SON) of the rat. Laser capture microdissection and qRT-PCR was used to demonstrate BDNF and TrkB gene expression in vasopressin SON neurones. Immunohistochemistry confirmed BDNF staining in vasopressin neurones, while staining for phosphorylated TrkB was increased following water deprivation. Western Blot analysis of brain punches containing the SON revealed that tyrosine phosphorylation of TrkB (pTrkBY515), serine phosphorylation of NR1 (pNR1S866 or pNR1) and tyrosine phosphorylation of NR2B subunits (pNR2BY1472 or pNR2B) were significantly increased in WD animals compared to control. Access to water for 2 h reduced pTrkBY515 content to control levels without affecting pNR1 or pNR2B. Four hours of rehydration was needed to reduce pNR1 and pNR2B to control. To test whether increased phosphorylation of TrkB in this study is mediated by BDNF, a group of animals were instrumented with right SON cannula coupled to mini-osmotic pumps filled with vehicle or TrkB-Fc fusion protein which prevents BDNF binding to TrkB. In the left SON contralateral to the cannula, TrkB phosphorylation was significantly enhanced following WD. Separate analysis of the right SON, which received TrkB-Fc, showed that the TrkB receptor phosphorylation following WD was significantly attenuated. While increased pNR1S866 following WD was not affected by local infusion of TrkB-Fc, pNR2BY1472 was significantly reduced. Co-immunoprecipitation revealed an increased physical interaction between Fyn kinase and NR2B and TrkB in the SON following water deprivation. Thus, activation of TrkB in the SON following WD may affect cellular excitability through the phosphorylation of NR2B subunits.