We have found that, during the early stages of cortical neurogenesis, both GABA and glutamate depolarize cells in the ventricular zone of rat embryonic neocortex. In the ventricular zone, glutamate acts on AMPA/kainate receptors, while GABA acts on GABAA receptors. GABA induces an inward current at resting membrane potentials, presumably owing to a high intracellular Cl- concentration maintained by furosemide-sensitive Cl- transport. GABA and glutamate also produce increases in intracellular Ca2+ in ventricular zone cells, in part through activation of voltage-gated Ca2+ channels. Furthermore, GABA and glutamate decrease the number of embryonic cortical cells synthesizing DNA. Depolarization with K+ similarly decreases DNA synthesis, suggesting that the neurotransmitters act via membrane depolarization. Applied alone, GABAA and AMPA/kainate receptor antagonists increase DNA synthesis, indicating that endogenously released amino acids influence neocortical progenitors in the cell cycle. These results demonstrate a novel role for amino acid neurotransmitters in regulating neocortical neurogenesis.

The behavioral and cognitive effects of nicotine suggest that nicotinic acetylcholine receptors (nAChRs) participate in central nervous system (CNS) function. Although nAChR subunit messenger RNA (mRNA) and nicotine binding sites are common in the brain, there is little evidence for synapses mediated by nAChRs in the CNS. To test whether, CNS nAChRs might modify rather than mediate transmission, the regulation of excitatory synaptic transmission by these receptors was examined. Nanomolar concentrations of nicotine enhanced both glutamatergic and cholinergic synaptic transmission by activation of presynaptic nAChRs that increased presynaptic [Ca2]i. Pharmacological and subunit deletion experiments reveal that these presynaptic nAChRs include the alpha 7 subunit. These findings reveal that CNS nAChRs enhance fast excitatory transmission, providing a likely mechanism for the complex behavioral effects of nicotine.

The period of hypoxia is an important priming event for the vascular dysfunction that accompanies reperfusion, with endothelial cells (ECs) and neutrophils (PMNs) playing a central role. We hypothesized that EC Weibel-Palade (WP) body exocytosis during the hypoxic/ischemic period during organ preservation permits brisk PMN recruitment into postischemic tissue, a process further amplified in an oxidant-rich milieu. Exposure of human umbilical vein ECs to a hypoxic environment (pO 2 ഠ 20 torr) stimulated release of von Willebrand factor (vWF), stored in EC WP bodies, as well as increased expression of the WP body-derived PMN adhesion molecule P-selectin at the EC surface. Increased binding of 111 In-labeled PMNs to hypoxic EC monolayers (compared with normoxic controls) was blocked with a blocking antibody to P-selectin, but was not affected by a nonblocking control antibody. Although increased P-selectin expression and vWF release were also noted during reoxygenation, hypoxia alone (even in the presence of antioxidants) was sufficient to increase WP body exocytosis. To determine the relevance of these observations to hypothermic cardiac preservation, during which the pO 2 within the cardiac vasculature declines to similarly low levels, experiments were performed in a rodent (rat and mouse) cardiac preservation/transplantation model. Immunodepletion of recipient PMNs or administration of a blocking anti-Pselectin antibody before transplantation resulted in reduced graft neutrophil infiltration and improved graft survival, compared with identically preserved hearts transplanted into control recipients. To establish the important role of endothelial P-selectin expression on the donor vasculature, murine cardiac transplants were performed using homozygous P-selectin deficient and wild-type control donor hearts flushed free of blood/platelets before preservation/transplantation. P-selectin-null hearts transplanted into wildtype recipients demonstrated a marked (13-fold) reduction in graft neutrophil infiltration and increased graft survival compared with wild-type hearts transplanted into wild-type recipients. To determine whether coronary endothelial WP exocytosis may occur during cardiac preservation in humans, the release of vWF into the coronary sinus (CS) was measured in 32 patients during open heart surgery. CS samples obtained at the start and conclusion of the ischemic period demonstrated an increase in CS vWF antigen (by ELISA) consisting of predominantly high molecular weight multimers (by immunoelectrophoresis). These data suggest that EC WP exocytosis occurs during hypothermic cardiac preservation, priming the vasculature to recruit PMNs rapidly during reperfusion. ( J. Clin. Invest. 1996. 97:493-500.)

Alterations in serotonin (5-hydroxytriptamine, 5-HT), norepinephrine, and ␥-aminobutyric acid have been linked to the pathophysiology of anxiety and depression, and medications that modulate these neurotransmitters are widely used to treat mood disorders. Recently, the neuropeptide substance P (SP) and its receptor, the neurokinin 1 receptor (NK1R), have been proposed as possible targets for new antidepressant and anxiolytic therapies. However, animal and human studies have so far failed to provide a clear consensus on the role of SP in the modulation of emotional states. Here we show that both genetic disruption and acute pharmacological blockade of the NK1R in mice result in a marked reduction of anxiety and stress-related responses. These behavioral changes are paralleled by an increase in the firing rate of 5-HT neurons in the dorsal raphe nucleus, a major source of serotonergic input to the forebrain. NK1R disruption also results in a selective desensitization of 5-HT1A inhibitory autoreceptors, which resembles the effect of sustained antidepressant treatment. Together these results indicate that the SP system powerfully modulates anxiety and suggest that this effect is at least in part mediated by changes in the 5-HT system. S ubstance P (SP) and its receptor, the neurokinin 1 receptor (NK1R), participate in the neural processing of a range of noxious and stressful stimuli. In the spinal cord, SP contributes to nociception, and disruption of the NK1R decreases or ablates the late-phase response to peripheral injury (1-3). In the brainstem, SP modulates emesis, which can be decreased in animals and humans by NK1R antagonists (4). Peripheral inflammation in a variety of structures including gut, joints, and cutaneous tissue also partly depends on SP release and NK1R activation (5).Recently, SP also has been implicated in the modulation of stress responses, mood, and anxiety, but its exact role remains unclear. Localized administration of SP in the central nervous system may produce anxiogenic or anxiolytic responses, depending on the animal species and location of injection (6-8). Conflicting results also have been obtained from the use of NK1R antagonists, and issues of drug access and species specificity have further clouded the roles of SP in stress-related behaviors (9). Interestingly, in humans, an NK1R antagonist has been reported to be an effective antidepressant and anxiolytic, although this result has not been replicated (10). It has been proposed that the antidepressant activity of NK1R antagonism may be independent of serotonergic and noradrenergic pathways and thus may represent an alternative therapeutic strategy for the treatment of mood disorders (10).In the present study we used genetic disruption and pharmacological blockade of the NK1R to examine the roles of SP in the generation of anxiety-related behaviors in mice. Genetic disruption of NK1R function markedly reduced anxiety-related behaviors in the elevated plus maze (EPM), novelty suppressed feeding (NSF), and maternal separation paradi...

Vasopressin is a uniquely effective pressor in the irreversible phase of hemorrhagic shock unresponsive to volume replacement and catecholamine vasopressors. Vasopressin deficiency may contribute to the pathogenesis of this condition.

1. Expression of receptors to extracellular calcium enables parafollicular cells of the thyroid gland (PF cells) to release calcitonin (CT) and serotonin (5_HT) in response to increased external Ca¥. Recently, a calcium-sensing receptor (CaR), similar to the G protein-coupled receptor for external Ca¥ cloned from parathyroid gland, was shown to be expressed in PF cells. Using a highly purified preparation of sheep PF cells, we have examined the electrical and biochemical processes coupling CaR activation to hormone release. cation is Ca¥, indicating that influx via the cation conductance is another source of the increases in [Ca¥]é. 6. Extracellular Ca¥ stimulates 5_HT release with an ECÛÑ of 1·5 mÒ. Nimodipine blocks 90% of the Cafi-induced 5_HT release, while other inhibitors of voltage-gated calcium channels had no effect. These data support an important role for L-type Ca¥ channels in CaR-induced hormone secretion. Although earlier studies indicate that high [Ca¥]ï induces release of Ca¥ from intracellular stores, thapsigargin-induced depletion of these stores did not affect secretion from these cells, indicating that Ca¥ influx is necessary and sufficient for the Cafi-induced 5_HT secretion. 7. Inhibition of protein kinase C (PKC) using chelerythrine, staurosporine, or calphostin C inhibited Cafi-induced 5_HT release by 50% while phorbol ester-induced 5_HT secretion was completely inhibited. Thus, PKC is an important component of the pathway linking CaR activation to hormone release. However, another as yet unknown second messenger also contributes to this pathway. 8. We tested the contribution of two different phospholipases to the CaR responses to determine the source of the PKC activator diacylglycerol (DAG). Selective inhibition of phosphatidylinositol-specific phospholipase C (PI-PLC) with U73122 had no effect on the response to elevated [Ca¥]ï. However, pretreatment with D609, a selective inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC), inhibited Ca¥-induced 5_HT release to 50% of control indicating that phosphatidylcholine is a likely source of DAG in the response of PF cells to elevated [Ca¥]ï.

1. A Ca(2+)‐dependent interaction between non‐NMDA and NMDA receptors was studied in embryonic rat dorsal horn neurons grown in tissue culture using perforated‐patch recording. Specifically, non‐NMDA receptors were found to induce reversible inhibition of NMDA receptors in a manner dependent on the presence of extracellular Ca2+. 2. Non‐NMDA receptor‐induced inhibition of NMDA receptors was mediated by the elevation of intracellular Ca2+ concentration produced by Ca2+ entry through a subpopulation of alpha‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate (AMPA) non‐NMDA receptors. Furthermore, Ca2+ entry through the AMPA channels alone is sufficient for desensitization of NMDA channels to occur. 3. Imaging of neuritic sites of Ca2+ revealed that Ca(2+)‐permeable AMPA channels are often co‐localized with NMDA channels on dorsal horn neurons, indicating that the Ca(2+)‐mediated interaction between receptors may occur within small dendritic domains. 4. The ability of Ca(2+)‐permeable AMPA channels to inhibit adjacent NMDA channels may contribute to the postsynaptic integration of excitatory input.

1. We used microfluorimetric measurement of [Ca2+]i to identify substance P-sensitive cells acutely isolated from the dorsal horn of neonatal rats. We then used morphological, physiological, and immunocytochemical criteria to delineate two distinct populations of substance P-sensitive dorsal horn cells. 2. One population of cells with small-diameter cell bodies and many fine processes responds to substance P by releasing Ca2+ from internal stores. Many of these cells express the O4 surface antigen, and are thus likely to be glial cells, probably from the oligodendrocyte lineage. None of the cells with glial attributes respond to N-methyl-D-aspartate (NMDA), providing further evidence that they are nonneuronal. 3. In a second population of dorsal horn cells, substance P elevates [Ca2+]i by promoting Ca2+ entry. This class of cells is morphologically distinct from substance P-sensitive glial cells in that it exhibits large-diameter cell bodies, has smooth tapering processes, and is sensitive to NMDA. This second class of cells is therefore likely to consist of neurons. 4. Consistent with the identification of different mechanisms of Ca2+ elevation in the two cell types, the kinetics of the substance P-evoked release of Ca2+ in glial cells is very different than the kinetics of the Ca(2+)-entry response evoked in neurons. The glial cell response had a rapid average rate of rise (mean = 260 +/- 105 nM/s) and relatively brief duration (mean = 7.6 +/- 2.2 s) whereas the neuronal response had a much slower rate of rise (mean = 10 +/- 9 nM/s) with a much longer duration.(ABSTRACT TRUNCATED AT 250 WORDS)