Francis M. Sessler

Noradrenergic potentiation of excitatory transmitter action in cerebrocortical slices: evidence for mediation by an α1 receptor-linked second messenger pathway

Considerable evidence from intact, anesthetized preparations suggests that norepinephrine (NE) can modulate the efficacy of synaptic transmission within local circuits of the mammalian neocortex; i.e. both iontophoretic application of NE and activation of the coeruleocortical pathway are capable of facilitating cortical neuronal responses to non-noradrenergic synaptic inputs and putative transmitter agents. In the present study, the effects of NE on somatosensory cortical neuronal responses to putative excitatory transmitters were characterized using in vitro tissue slice preparations. Somatosensory unit responses to iontophoretic pulses of acetylcholine (ACh) or glutamate (Glu) (10-60 nA; 5-25 s duration) were examined before, during and after a period of continuous NE (1-35 nA; 4-25 min duration) microiontophoresis. Quantitative analysis of per-event histograms indicated that both Glu- and ACh-evoked excitatory discharges were routinely (Glu 94%, n = 54; ACh 67%, n = 9) potentiated above control levels during NE administration. In 8 cells, NE revealed robust excitatory discharges to otherwise subthreshold iontophoretic doses of Glu. The alpha-specific agonist, phenylephrine, mimicked (n = 3), NE-induced potentiation of Glu-evoked discharges whereas the alpha antagonist phentolamine blocked (n = 5) enhancement of these responses. Moreover, activation of protein kinase C by iontophoretic application of phorbol 12,13-diacetate (5-15 nA, n = 4) mimicked the potentiating actions of NE on Glu-evoked excitatory responses. Results from other experiments further indicated that these facilitating actions of NE on Glu-evoked responses do not involve beta receptor activation or intracellular increases in cyclic AMP. In summary, these results demonstrate that NE can facilitate cortical neuronal responses to threshold and subthreshold level applications of putative excitatory transmitter agents. Moreover, it appears that, unlike noradrenergic facilitating influences on GABA-induced inhibition, these actions are mediated by an alpha adrenoceptor mechanism which may be linked to intracellular activation of protein kinase C. Overall, these findings reinforce the idea that noradrenergic modulatory actions on excitatory and inhibitory neuronal responses may involve the activation of separate receptor-linked second messenger systems.

Neurotrophic factor mRNA expression in dentate gyrus is increased following in vivo stimulation of the angular bundle.

Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are two structurally-related neurotrophins synthesized in dentate gyrus granule cells and pyramidal neurons of the hippocampal formation. These neurons receive excitatory glutamatergic afferents from the entorhinal cortex via the angular bundle/perforant path. In the present study, we tested whether electrophysiological stimulation of this glutamatergic pathway modifies NGF or BDNF messenger RNA (mRNA) expression in vivo. Within hours following brief trains of high frequency angular bundle stimulation, the levels of mRNA encoding both neurotrophins were increased exclusively in granule cells of the ipsilateral dentate gyrus. The increase in neurotrophic factor mRNA expression was found to be mediated through the N-methyl-D-aspartate (NMDA) glutamate receptor subtype, and occurred in the absence of seizure. These findings provide evidence that neurotrophic factor mRNA levels in the hippocampal formation are increased by direct activation of excitatory afferents originating in the entorhinal cortex. We suggest that the function of some neurotrophin-responsive neuronal populations may depend upon the integrity and activity of neurons in the entorhinal cortex, a population of neurons reported to be compromised in patients with Alzheimers disease.

Differential modulatory effects of norepinephrine on synaptically driven responses of layer V barrel field cortical neurons.

The effects of norepinephrine (NE) and the alpha-1 agonist phenylephrine (PE) on synaptically evoked responses of electrophysiologically identified pyramidal neurons in layer V of rat somatosensory cortex were studied in brain slices using intracellular recording techniques. When added to the bathing medium NE (10 microM) tended to increase the synaptic responsiveness of regular spiking neurons and decrease the responsiveness of intrinsic burst neurons. NE had mixed effects on layer V cells which were characterized as intermediate types between regular spiking and intrinsic burst neurons. PE exerted a similar spectrum of actions on layer V cortical neurons. For both adrenergic agents the greatest facilitating effect was observed on responses to low intensity synaptic stimulation. These results suggest that NE exerts different modulatory actions on different electrophysiologically-defined classes of layer V sensory cortical neurons.

Regulation of nerve growth factor mRNA in the hippocampal formation : effects of N-methyl-D-aspartate receptor activation

In the hippocampal formation, nerve growth factor (NGF) is produced in granule cells of the dentate gyrus and a few pyramidal cells of Ammons horn. Both neuronal populations express N-methyl-D-aspartate (NMDA) receptors and receive putative glutamatergic afferents originating in the entorhinal cortex and projecting via the perforant path. We report in this study that intra-hippocampal or intraventricular injections of NMDA increase NGF mRNA levels in dentate gyrus granule cells as determined using in situ hybridization histochemistry and a solution hybridization assay. NGF mRNA induction is detected within 2 h following NMDA treatment and returns to control levels within 24 h. This NMDA effect is dose-dependent and blocked by pretreatment with 2-amino-5-phosphonopentanoic acid, a competitive NMDA antagonist. Finally, the induction of NGF mRNA is observed in the absence of detectable neurotoxicity or seizure activity. We postulate that normal physiological events associated with the activation of hippocampal NMDA receptors may regulate mRNA expression of this neurotrophic factor.

Noradrenergic enhancement of GABA-induced input resistance changes in layer V regular spiking pyramidal neurons of rat somatosensory cortex

Previous in vivo studies have shown that microiontophoretic application of norepinephrine (NE) and isoproterenol (ISO) can enhance gamma-aminobutyric acid (GABA)-induced depressant responses of rat somatosensory cortical neurons. In the present investigation we have examined the transmembrane electrophysiological events which are associated with interactions between NE and GABA in layer V pyramidal neurons of rat barrel field cortex. Intracellular recordings were made from electrophysiologically identified cells in a superfused cortical tissue slice preparation before, during and after bath or microdrop application of GABA, NE and ISO, alone or in combination. GABA application produced a small depolarization from resting membrane potential associated with a reduction (22%) in input resistance. NE and ISO (10-100 microM) also produced in some cases small membrane depolarizations (1-4 mV) but little concomitant changes in input resistance. Simultaneous application of NE with GABA potentiated amino acid-induced changes in input resistance in 4 cases and antagonized (n = 4) or had no effect (n = 4) on GABA-associated membrane events in 8 other cases. When the alpha-blocker, phentolamine (20 microM), was added to the medium, NE-induced enhancement of the GABA response was observed in 3 of 5 cases (60%), suggesting both, a beta-adrenergic mediation and a possible alpha-receptor masking of this noradrenergic-potentiating action. Consistent with this interpretation was the finding that the beta-agonist, ISO (10-100 microM), produced net increases in GABA-induced input resistance changes in 64% of cases tested (9 of 14). The potentiating effect of NE and ISO was mimicked by the adenyl cyclase activator, forskolin (n = 2), and a membrane permeant analog of cyclic-AMP, 8-bromo-cyclic AMP (n = 3); and could also be demonstrated when the GABAA agonist muscimol (0.5-1 microM) was substituted for GABA. The reversal potential for GABA and GABA + NE remained the same. These findings suggest that previous demonstrations of NE-potentiating effects on GABA inhibition may be mediated by beta-receptor/cyclic-AMP-linked actions on mechanisms which regulate GABAA receptor-induced membrane conductance changes.

Glial localization of adenylate-cyclase-coupled β-adrenoreceptors in rat forebrain slices

Abstract Fluorocitrate (FC), a selective inhibitor of glial cell respiration, was used to estimate the extent to which glial cells contain adenylate cyclase-coupled β-adrenoreceptors in rat brain slices. The drug blocked 75–95% of the elevation of cyclic AMP caused by the β-agonist, isoproterenol, in the 4 forebrain regions sampled (frontal and parietal cortex, caudate nucleus, olfactory tubercle). Intracellular recording of neurons in the treated slices confirmed that they were unaffected by FC. Treatment with the neurotoxin, kainic acid, eliminated all electrophysiological activity but did not affect the cAMP response. The results indicate that glial cells contain the preponderance of adenylate-cyclase-coupled β-adrenoreceptors in slices of the rat forebrain and may constitute an important target of the central noradrenergic system in vivo.

Neurotrophic factor mRNA expression in dentate gyrus is increased following angular bundle transection

In the central nervous system, the highest levels of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) mRNA are found in the hippocampal formation. In the present study, we report that a unilateral transection of the angular bundle, which relays cortical information via the entorhinal cortex to the hippocampal formation, increases NGF and BDNF mRNA in the ipsilateral dentate gyrus. Within 4 hours following transection, the hybridization signal for NGF and BDNF mRNA increases in stratum granulosum 3- and 5-fold, respectively, compared to control levels. This lesion-induced increase of both mRNA returns to control levels within 24 hours and is maintained for at least 5 days. The induction is not prevented by pretreatment with AP-5, CNQX, or cholinergic denervation due to transection of the fimbria-fornix. Finally, the induction of neurotrophin mRNA is preceded by an increase in c-fos mRNA. These results provide evidence that transection of the cortical input to the hippocampal formation upregulates NGF and BDNF mRNA selectively in stratum granulosum. We suggest that the increased expression of NGF and BDNF mRNA may be an early step in the synaptic rearrangement of neurotrophin responsive cholinergic afferents observed following damage to the entorhinal cortex.

Second messenger-mediated actions of norepinephrine on target neurons in central circuits: a new perspective on intracellular mechanisms and functional consequences

Ever since the initial demonstration of a widespread distribution of noradrenergic fibers to functionally diverse regions of the mammalian forebrain, there has been considerable interest in determining the electrophysiological effects of norepinephrine (NE) on individual neurons within these target areas. While early studies showed that NE could directly inhibit cell firing via increased intracellular levels of cyclic AMP, more recent work has revealed a spectrum of noradrenergic actions, which are more accurately characterized as neuromodulatory. More specifically, numerous experimental conditions have been described where NE at levels subthreshold for producing direct depressant effects on spontaneous firing can facilitate neuronal responses to both excitatory and inhibitory synaptic stimuli. The goal of this report is to review recent evidence which suggests that the various modulatory actions of NE on central neurons result from the activation of different adrenoceptor-linked second messenger systems. In particular, we have focused on the candidate signal transduction mechanisms that may underlie NEs ability to augment cerebellar and cortical neuronal responsiveness to GABAergic synaptic inputs. The consequences of such NE-induced changes in synaptic efficacy are considered not only with respect to their influences on feature extraction properties of individual sensory cortical neurons but also with regard to the potential impact such actions would have on the signal processing capabilities of a network of noradrenergically innervated cortical cells.

Electrophysiological actions of norepinephrine in rat lateral hypothalamus. I. Norepinephrine-induced modulation of LH neuronal responsiveness to afferent synaptic inputs and putative neurotransmitters

The present studies were conducted as part of an ongoing investigation of the effects of norepinephrine (NE) in neuronal circuits of the mammalian brain. In this report, we describe noradrenergic actions in the lateral hypothalamus (LH), an area which has been implicated in the central integration of cardiovascular regulatory mechanisms, fluid balance and ingestive behaviors. Microiontophoretically applied NE was interacted with extracellularly recorded responses of LH neurons to iontophoretically applied putative neurotransmitters gamma-aminobutyric acid (GABA), acetylcholine (ACh) and glutamate (Glu); and activation of known input pathways from the reticular thalamus (RT) and the lateral preoptic area (LPO). Peri-event histograms of cell responses were computed before, during and after NE microiontophoresis (5-50 nA) and used to quantitatively evaluate monoamine-induced effects on spontaneous and stimulus evoked activity of LH neurons. In 16 of 23 LH neurons, RT-stimulus-induced inhibition was markedly prolonged from a mean of 28.3 +/- 4.8 ms to 44.7 +/- 5.2 ms, during iontophoretic application of NE. In 22 of 38 LH cells, LPO-stimulus-induced excitatory responses were enhanced above control levels during NE administration. In further tests, inhibitory responses of LH cells to iontophoretic pulses of GABA were potentiated during NE administration in 69% (24 of 35) of the cases tested. ACh-induced excitation was potentiated in 9 of 21 cells. In 4 of these cases, otherwise subthreshold doses of ACh caused marked increases in cell firing during the period of NE administration. By contrast, Glu-evoked excitation was antagonized by NE iontophoresis in 65.5% (17 of 26) of LH cells tested. These findings indicate that, as in other noradrenergic target regions of the CNS, NE can facilitate synaptically mediated responses of LH neurons. Taken together these observations suggest that NE may play an important regulatory role in the synaptic transfer of information within LH circuits, and consequently exert considerable influence over the homeostatic functions mediated by this structure.

New perspectives on the functional organization and postsynaptic influences of the locus ceruleus efferent projection system.

Publisher Summary This chapter summarizes recent efforts on the design of anatomical and physiological experiments that are capable of bridging the gap between cellular-membrane studies of locus ceruleus-norepinephrine (LC-NE) attributes and more global hypotheses regarding the impact of the LC projection system on sensory neural network function. Recent studies using fluorescent retrograde tracers have shown that LC efferents to target structures along the trigeminal somatosensory pathway in rat exhibit an orderly projection with respect to the crossed trajectory of this ascending sensory system. It has been also found that LC projection neurons to somatosensory structures are organized into overlapping subsets within the nucleus, thus raising the possibility that a significant proportion of these cells may project to multiple, common somatosensory targets via axon collaterals. These studies show that the LC exhibits an internal organization and efferent projection pattern that is consistent with the idea that LC outputs can selectively and coordinately modulate the flow of somatosensory information through forebrain circuits. These findings indicate that NE can augment membrane responses to GABA via a β -receptor-linked/ cyclic adenosine monophosphate (cAMP) cascade of events. As such, these noradrenergic influences on putative transmitter-induced conductance changes are consistent with previously observed effects of iontophoretic NE on single-unit responses to GABA in anesthetized and awake, behaving animals. The results reported here continue to support and refine the idea that the LC efferent system exhibits a high degree of anatomical and physiological specificity with respect to its potential impact on target neuronal circuits. Overall, these advances in the understanding of the organizational principles and physiological specificity of the LC-NE efferent system provide new insights into the role of this monoamine-containing brain-stem nucleus in behavior.