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This study presents a novel mathematical modeling framework that is uniquely suited to investigating the structure and dynamics of the sleep-wake regulatory network in the brain stem and hypothalamus. It is based on a population firing rate model formalism that is modified to explicitly include concentration levels of neurotransmitters released to postsynaptic populations. Using this framework, interactions among primary brain stem and hypothalamic neuronal nuclei involved in rat sleep-wake regulation are modeled. The model network captures realistic rat polyphasic sleep-wake behavior consisting of wake, rapid eye movement (REM) sleep, and non-REM (NREM) sleep states. Network dynamics include a cyclic pattern of NREM sleep, REM sleep, and wake states that is disrupted by simulated variability of neurotransmitter release and external noise to the network. Explicit modeling of neurotransmitter concentrations allows for simulations of microinjections of neurotransmitter agonists and antagonists into a key wake-promoting population, the locus coeruleus (LC). Effects of these simulated microinjections on sleep-wake states are tracked and compared with experimental observations. Agonist/antagonist pairs, which are presumed to have opposing effects on LC activity, do not generally induce opposing effects on sleep-wake patterning because of multiple mechanisms for LC activation in the network. Also, different agents, which are presumed to have parallel effects on LC activity, do not induce parallel effects on sleep-wake patterning because of differences in the state dependence or independence of agonist and antagonist action. These simulation results highlight the utility of formal mathematical modeling for constraining conceptual models of the sleep-wake regulatory network.

Light-evoked currents in depolarizing and hyperpolarizing bipolar cells (DBCs and HBCs) were recorded under voltage-clamp conditions in living retinal slices of the larval tiger salamander. Responses to illumination at the center of the DBCs' and HBCs' receptive fields were mediated by two postsynaptic currents: DeltaI(C), a glutamate-gated cation current with a reversal potential near 0 mV, and DeltaI(Cl), a chloride current with a reversal potential near -60 mV. In DBCs DeltaI(C) was suppressed by L-2-amino-4-phosphonobutyric acid (L-AP4), and in HBCs it was suppressed by 6,7-dinitroquinoxaline-2,3-dione (DNQX). In both DBCs and HBCs DeltaI(Cl) was suppressed by imidazole-4-acetic acid (I4AA), a GABA receptor agonist and GABA(C) receptor antagonist. In all DBCs and HBCs examined, 10 microM I4AA eliminated DeltaI(Cl) and the light-evoked current became predominately mediated by DeltaI(C). The addition of 20 microM L-AP4 to the DBCs or 50 microM DNQX to HBCs completely abolished DeltaI(C). Focal application of glutamate at the inner plexiform layer elicited chloride currents in bipolar cells by depolarizing amacrine cells that release GABA at synapses on bipolar cell axon terminals, and such glutamate-induced chloride currents in DBCs and HBCs could be reversibly blocked by 10 microM I4AA. These experiments suggest that the light-evoked, I4AA-sensitive chloride currents (DeltaI(Cl)) in DBCs and HBCs are mediated by narrow field GABAergic amacrine cells that activate GABA(C) receptors on bipolar cell axon terminals. Picrotoxin (200 microM) or (1,2,5,6-tetrahydropyridine-4yl) methyphosphinic acid (TPMPA) (2 other GABA(C) receptor antagonists) did not block (but enhanced and broadened) the light-evoked DeltaI(Cl), although they decreased the chloride current induced by puff application of GABA or glutamate. The light response of narrow field amacrine cells were not affected by I4AA, but were substantially enhanced and broadened by picrotoxin. These results suggest that there are at least two types of GABA(C) receptors in bipolar cells: one exhibits stronger I4AA sensitivity than the other, but both can be partially blocked by picrotoxin. The GABA receptors in narrow field amacrine cells are I4AA insensitive and picrotoxin sensitive. The light-evoked DeltaI(Cl) in bipolar cells are mediated by the more strongly I4AA-sensitive GABA(C) receptors. Picrotoxin, although acting as a partial GABA(C) receptor antagonist in bipolar cells, does not suppress DeltaI(Cl) because its presynaptic effects on amacrine cell light responses override its antagonistic postsynaptic actions.

Nucleus HVc of the songbird is a distinct forebrain region that is essential for song production and shows selective responses to complex auditory stimuli. Two neuronal populations within HVc give rise to its efferent projections. One projection, to the robust nucleus of the archistriatum (RA), serves as the primary motor pathway for song production, and can also carry auditory information to RA. The other projection of HVc begins a pathway through the anterior forebrain, (area X --> medial portion of the dorsolateral nucleus of the thalamus (DLM) --> lateral portion of the magnocellular nucleus of the anterior neostriatum (L-MAN) --> RA) that is crucial for song learning but, although active during singing, is not essential for adult song production. To test whether these different projection neuron classes have different functional properties, we recorded intracellularly from neurons in nucleus HVc in brain slices. We observed at least three classes of neuron based on intrinsic physiological and pharmacological properties as well as on synaptic inputs. We also examined the morphological properties of the cells by filling recorded neurons with neurobiotin. The different physiological cell types correspond to separate populations based on their soma size, dendritic extent, and axonal projection. Thus HVc neurons projecting to area X have large somata, show little spike-frequency adaptation, a hyperpolarizing response to the metabotropic glutamate receptor (mGluR) agonist (1S,3R)-trans-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD), and exhibit a slow inhibitory postsynaptic potential (IPSP) following tetanic stimulation. Those HVc neurons projecting to motor nucleus RA have smaller somata, show strong accommodation, are not consistently hyperpolarized by ACPD, and exhibit no slow IPSP. A third, rarely recorded class of neurons fire in a sustained fashion at very high-frequency and may be interneurons. Thus the neuronal classes within HVc have different functional properties, which may be important for carrying specific information to their postsynaptic targets.

The interaction between neurohumoral agonists and glucose to stimulate phosphoinositide (PI)-specific phospholipase C (PLC) and insulin release was examined. In freshly isolated rat islets, maximal glucose (40 mM), cholecystokinin (CCK; 300 nM), or carbachol (CCh; 1 mM) stimulated PI hydrolysis 6.5-, 9.8-, and 5.7-fold, respectively, above basal. The combination of glucose and CCK or of glucose and CCh, but not of CCK and CCh, synergistically increased PI hydrolysis 23.2- and 21.6-fold, respectively, indicating that these secretagogues activate PLC by distinct pathways and that there is an interaction between them. This synergy was maximal at physiological concentrations of stimulatory glucose (8-10 mM) and was paralleled by a marked synergistic stimulation of insulin secretion. The enhanced PI response was partially Ca2+ dependent and may involve the activation of distinct isozymes of PLC, which we identify in islets. These studies demonstrate for the first time a unique and highly sensitive synergistic interaction between glucose and neurohumoral agonists to stimulate PI hydrolysis, and they suggest that enhanced PI hydrolysis is important in the potentiation of glucose- and neurohumoral-stimulated insulin secretion.

To test the hypothesis that the distension-induced esophageal peristaltic reflex involves a polysynaptic pathway, a triple-chamber organ bath was used to chemically isolate different regions of the opossum smooth muscle esophagus while leaving in continuity the intramural neuromuscular apparatus. Balloon distension in the oral chamber evoked membrane hyperpolarization in the aboral chamber, which was followed, on balloon deflation, by depolarization, spike burst, and circular smooth muscle contraction. This reflex was abolished by adding tetrodotoxin to any of the chambers. Addition of Ca(2+)-free 20 mM Mg2+ Krebs solution (to block synaptic transmission) to the intermediate chamber did not affect the descending peristaltic reflex. However, Ca(2+)-free 20 mM Mg2+ Krebs solution markedly attenuated the reflex when placed in either the oral or the aboral chamber. Furthermore, the nitric oxide synthase inhibitor N omega-nitro-L-arginine methyl ester, when placed in the aboral chamber, abolished the reflex. Other putative neurotransmitter antagonists were without effect when placed in any chamber. These studies suggest that the intramural neural pathway that mediates the descending peristaltic reflex in the opossum esophagus is not polysynaptic, but rather involves long descending neurons that depend on nitric oxide as a final mediator.