Thesis (Ph. D.)--University of Virginia, 2007. / Includes bibliographical references. Also available online through Digital Dissertations.

Diss. (sammanfattning) Umeå : Umeå universitet, 2007. / Härtill 7 uppsatser.

Thesis (Ph. D.)--University of Washington, 1997. / Vita. Includes bibliographical references (leaves [110]-127).

A novel solubilising agent (0.l% sodium cholate-lM NaCl) has been developed which will solubilise 10-30% of muscarinic cholinergic receptors from bovine caudate nucleus. Using the muscarinic antagonist quinuclidinyl benzilate (QNB), a single saturable binding component was found with an equilibrium constant of ZOOpM, approximately 10-fold higher than the membrane receptor and 4-fold higher than the ratio k_-l/k_l determined kinetically in the soluble material. This latter difference may indicate that the binding of QNB to the solubilised receptor is not a simple second-order process. Inhibition constants for a variety of muscarinic agonists and antagonists were 10 to 20-fold higher than in the membrane state and non-muscarinic ligands were without effect. The decrease in affinity was shown to be due to the presence of high salt. Evidence was presented that the apparent increase in Hill coefficient for muscarinic agonist binding to soluble material was not due to a differential solubilisation of muscarinic receptors or to a conformational change of high to low affinity agonist sites during the solubilisation. Instead the Hill coefficient of the soluble material decreased as the percentage of total binding sites solubilised increased. The stability of receptor binding at different temperatures was shown to be dependent on the protein: cholate (w/w) ratio. Results from gel filtration, affinity chromotography and immunization studies are also reported. The results of this thesis are discussed in the light of the possible importance of phospholipids for receptor activity.

No description available.

Theories of the neurobiological basis of Attention-Deficit/Hyperactivity Disorder (ADHD) have largely focused on dysregulation of central dopaminergic function. However, other neurotransmitter systems may be implicated in specific cognitive deficits in ADHD. Interest in the potential involvement of nicotinic cholinergic systems in ADHD has arisen in part from the observation that adolescents and adults with ADHD smoke cigarettes at significantly higher rates than people without this disorder. In addition, several studies report that nicotine alleviates ADHD symptoms, and recent neuro-genetics studies indicate that cholinergic systems may be altered in persons with ADHD. In this review, we describe the evidence for a role of central nicotinic cholinergic systems in cognitive deficits in ADHD. We also propose mechanisms by which alterations in cholinergic function may contribute directly and/or indirectly to these deficits. Finally, we identify specific paradigms and models to guide future investigations into the specific involvement of nicotinic cholinergic systems in ADHD, possibly leading to the development of more effective pharmacotherapies for ADHD.

Rapid eye movement sleep decreases between 10 and 30 days postnatally in the rat. The pedunculopontine nucleus is known to modulate waking and rapid eye movement sleep, and pedunculopontine nucleus neurons are thought to be hyperpolarized by noradrenergic input from the locus coeruleus. The goal of the study was to investigate the possibility that a change in α-2 adrenergic inhibition of pedunculopontine nucleus cells during this period could explain at least part of the developmental decrease in rapid eye movement sleep. We, therefore, recorded intracellularly in 12-21 day rat brainstem slices maintained in oxygenated artificial cerebrospinal fluid. Putative cholinergic vs. non-cholinergic pedunculopontine nucleus neurons were identified using nicotinamide adenine dinucleotide phosphate diaphorase histochemistry and intracellular injection of neurobiotin (Texas Red immunocytochemistry). Pedunculopontine nucleus neurons also were identified by intrinsic membrane properties, type I (low threshold spike), type II (A) and type III (A+low threshold spike), as previously described. Clonidine (20 μM) hyperpolarized most cholinergic and non-cholinergic pedunculopontine nucleus cells. This hyperpolarization decreased significantly in amplitude (mean±S.E.) from -6.8±1.0 mV at 12-13 days, to -3.0±0.7 mV at 20-21 days. However, much of these early effects (12-15 days) were indirect such that direct effects (tested following sodium channel blockade with tetrodotoxin (0.3 μM)) resulted in hyperpolarization averaging -3.4±0.5 mV, similar to that evident at 16-21 days. Non-cholinergic cells were less hyperpolarized than cholinergic cells at 12-13 days (-1.6±0.3 mV), but equally hyperpolarized at 20-21 days (-3.3±1.3 mV). In those cells tested, hyperpolarization was blocked by yohimbine, an α-2 adrenergic receptor antagonist (1.5 μM). These results suggest that the α-2 adrenergic receptor on cholinergic pedunculopontine nucleus neurons activated by clonidine may play only a modest role, if any, in the developmental decrease in rapid eye movement sleep. Clonidine blocked or reduced the hyperpolarization-activated inward cation conductance, so that its effects on the firing rate of a specific population of pedunculopontine nucleus neurons could be significant. In conclusion, the α-2 adrenergic input to pedunculopontine nucleus neurons appears to consistently modulate the firing rate of cholinergic and non-cholinergic pedunculopontine nucleus neurons, with important effects on the regulation of sleep-wake states.

The striatum is the initial input nuclei of the basal ganglia, and it serves as an integral processing center for action selection and sensorimotor learning. Glutamatergic projections from the cortex and thalamus converge with dense dopaminergic axons from the midbrain to provide the primary inputs to the striatum. Striatal output is then relayed to downstream basal ganglia nuclei by GABAergic medium – sized spiny neurons, which comprise at least 95% of the population of neurons in the striatum. The remaining population of local circuit neurons is dedicated to regulating the activity of spiny projection neurons, and although spiny neurons form a weak lateral inhibitory network among themselves via local axon collaterals, feedforward modulation exerts more powerful control over spiny neuron excitability. Of the striatal interneurons, only one class is not GABAergic. These neurons are cholinergic and correspond to the tonically active neurons (TANs) recorded in vivo, which respond to specific environmental stimuli with a transient depression, or pause, of tonic firing. Striatal cholinergic interneurons account for less than 2 % of the striatal neuronal population, yet their axons form an extensive and complex network that permeates the entire striatum and significantly shapes striatal output by acting at numerous targets via varied receptor types. Indeed, the persistent level of ambient striatal acetylcholine as well as changes to that basal acetylcholine level underlie the major mechanisms of cholinergic signaling in the striatum, however regulation of this system by the local striatal microcircuitry is not well understood. This dissertation finds that activation of intrastriatal cholinergic fibers elicits polysynaptic GABAA inhibitory postsynaptic currents (IPSCs) in cholinergic interneurons recorded in brain slices. Excitation of striatal GABAergic neurons via nicotinic acetylcholine receptors (nAChRs) mediates this polysynaptic inhibition in a manner independent of dopamine. Moreover, activation of a single cholinergic interneuron is capable of eliciting polysynaptic GABAA IPSCs onto itself and nearby cholinergic interneurons. These findings provide an important insight into the striatal microcircuitry controlling cholinergic neuron excitability. / text

Neuromodulatory systems, such as the cholinergic basal forebrain (CBF), are promising targets of behavioral neuroscientific research because of the clear role(s) they play in fundamental neural and behavioral plasticity processes. Previous research suggests that the CBF is a promising target for augmenting post-stroke behavioral rehabilitation. Several studies were conducted to establish a novel paradigm for investigating and ameliorating post-stroke related motor deficits in a rodent model. The first two studies describe the invention and validation of a novel apparatus for conducting individualized rodent research in an environmentally and socially enriched context. The second study specifically investigated how this approach yields novel insights into post-stroke motor deficits. The third study describes how electrical stimulation of the CBF may improve poststroke motor rehabilitation. Together, these studies are expected to improve current stroke models, our understanding of how the CBF supports fundamental learning processes, and how to best manipulate the CBF to improve recovery from neurological injury.

Located in the mesopontine tegmentum, the pedunculopontine tegmental nucleus (PPTg) is comprised principally of glutamatergic, cholinergic and GABAergic neurons. In addition to being fully integrated into basal ganglia, PPTg projects to thalamus and motor output sites in the brainstem. Previous studies have shown a range of behavioural changes after PPTg manipulation. Prominent amongst these is an apparent deficit in the ability to learn the consequences of actions. PPTg is divisible into a posterior component (pPPTg) in receipt of rapid polymodal sensory input and projecting into VTA/SNc dopamine neurons and an anterior component (aPPTg) in receipt of basal ganglia outflow and projecting into SNc and lower brainstem structures. The research described here assesses the role of the pPPTg in instrumental learning. Using a contingency degradation paradigm, it was shown that inactivation of the pPPTg (by muscimol microinfusion) specifically blocked the updating of associations between actions and outcomes, without the affecting the ability to re-execute previously learned instrumental actions. Selective bilateral destruction of pPPTg cholinergic neurons (with the fusion toxin diphtheria toxin – urotensin II [Dtx-UII]) resulted in >90% loss of pPPTg cholinergic neurons. These lesions produced no detectable changes on any measured aspect of an instrumental learning task consisting of various fixed and variable ratio schedules of reinforcement and extinction. Subsequent experiments found that the same selective cholinergic pPPTg lesions also produced no changes in the locomotor response to nicotine or rate of nicotine sensitisation. These results are the first to demonstrate a brainstem role in action-outcome learning. Results support the view that PPTg performs a ‘first pass' analysis on incoming sensory data and interfaces salient aspects of this with appropriate basal ganglia and brainstem circuitry, with glutamatergic pPPTg projections sending an essential signal and cholinergic projections performing as part of a wider modulatory system.