sfn2011

Savage ST, Olson L, Mattsson A (2011) Alterations in gene expression following cortical cholinergic denervation in rats. Neuroscience 2011 Abstracts 790.01. Society for Neuroscience, Washington, DC.

Summary: Alterations in cholinergic signaling in the brain have been implicated as a contributing factor in the pathogenesis of schizophrenia. Altered function and expression of both nicotinic and muscarinic acetylcholine receptors have been reported in cortical and subcortical regions in post-mortem schizophrenic brains. Pharmacologically, dopamine-releasing compounds, such as amphetamine, can induce the psychotic symptoms in healthy volunteers and exacerbate the symptoms in schizophrenics. Furthermore, the NMDA receptor antagonist phencyclidine (PCP) induces both negative symptoms (such as social withdrawal) and cognitive deficits similar to those exhibited in schizophrenics. We have previously shown that cholinergic denervation of cortex cerebri by stereotaxic infusion of the immunotoxin 192 IgG-saporin in the nucleus basalis magnocellularis (nbm) in adult rats leads to an enhanced sensitivity to both amphetamine and PCP. The enhanced sensitivity to amphetamine, shown by a potentiated dopamine release in nucleus accumbens, along with a marked increase in locomotor activity in response to both amphetamine and PCP, suggested that the disruption of cortical cholinergic activity can lead to disturbances of glutamatergic and dopaminergic transmission. Furthermore, bilateral lesioning of nbm led to a decrease in active social interaction, as well as, impairment after an acute PCP challenge in a cognitive task (novel object recognition). To further evaluate the consequences of cortical cholinergic denervation, we are analyzing the possible changes in mRNA expression levels of selected genes in rats with unilateral removal of the cortical cholinergic innervation by 192 IgG-saporin injections into nbm following acute PCP administration. Our data indicate that the induction of c-fos mRNA expression in cortex in response to PCP administration is markedly reduced in cholinergically denervated animals as compared to controls. Other genes are under investigation to elucidate the interplay between the cholinergic, dopaminergic, and glutamatergic systems.

Related Products: 192-IgG-SAP (Cat. #IT-01)

Wiley RG, Lappi DA (2011) Selective activation of dorsal horn inhibitory interneurons produces anti-nociception. Neuroscience 2011 Abstracts 804.14. Society for Neuroscience, Washington, DC.

Summary: Intrathecal injections of the excitatory neuropeptide neurotensin are antinociceptive in rats. Lumbar intrathecal injections of the cytotoxic conjugate, neurotensin-saporin (NTS-sap), cause rats to engage in intense scratching, licking and biting of their hindquarters. This observation was interpreted as indicating the rats were experiencing discomfort presumably because NTS-sap selectively destroys nociceptive inhibitory interneurons expressing neurotensin receptors in the superficial dorsal horn of the spinal cord resulting in decreased inhibitory input to nociceptive projection neurons. Based on this finding, we made the excitatory conjugate, neurotensin-cholera toxin A subunit (NTS-CTA) which we hypothesized would tonically activate the same nociceptive inhibitory interneurons and produce anti-nociception/analgesia. Two separate groups of Long Evans hooded female rats were injected, under general anesthesia, with 500 ng of NTS-CTA, produced by Advanced Targeting Systems, San Diego, CA using temporarily positioned subarachnoid catheters which were removed after 15 mins. For the next 72-96 hours, rats showed: 1 – normal spontaneous behavior including grooming, ambulation, defecation and urination; 2 – decreased nocifensive responses on the hotplate at 44C – 47C; 3 – increased hindpaw mechanical withdrawal thresholds; and, 4 – prolonged tail flick response latencies. Systemic naloxone (0.8-2.0 mg/kg, s.c.) did not reverse the anti-nociceptive effect of NT-CTA. Hotplate responses completely returned to baseline within 7 days. These data are interpreted as showing that intrathecal NTS-CTA is reversibly anti-nociceptive by a naloxone-insensitive (non-opioid) mechanism. The likely mechanism of NTS-CTA action is hypothesized to involve tonic activation of NTS receptor-expressing inhibitory interneurons in the superficial dorsal horn of the spinal cord that increases inhibition of nociceptive projection neurons. This strategy may prove useful in treating intractable pain and may be generally useful in the study and manipulation of other populations of inhibitory (or excitatory) interneurons using various neuropeptide-CTA conjugates in such fields as epilepsy, learning and memory, etc. Ongoing work is aimed at identifying the neurons activated by NTS-CTA, testing NTS-CTA in operant pain tests, testing nociceptive effects of other neuropeptide-CTA conjugates and evaluating ways to produce more prolonged activation of the target neurons.

Related Products: Neurotensin-SAP (Cat. #IT-56), Neurotensin-CTA (Cat. #IT-60)

Saeed AW, Ribeiro-Da-Silva A (2011) Consequences of the ablation of non-peptidergic nociceptive fibers on neurokinin-1 receptor expression by spinal lamina I neurons. Neuroscience 2011 Abstracts 804.21. Society for Neuroscience, Washington, DC.

Summary: Spinal dorsal horn lamina I projection neurons expressing the neurokinin-1 receptor (NK-1r) are important in relaying pain-related information from the periphery to the brain. These lamina I neurons have been classified, based on their morphological and physiological properties, into three types: fusiform, multipolar and pyramidal. Of these cell types, pyramidal neurons seldom express the NK-1r and are non-nociceptive. Previously, our laboratory has demonstrated in a cuff model of chronic constriction injury a de novo expression of NK-1r by pyramidal neurons, starting at the same time as the mechanical allodynia. We have also observed a similar de novo expression of NK-1r by pyramidal neurons in an animal model of arthritis. In the current study, we investigated whether the cytotoxic ablation of the non-peptidergic, isolectin B4 (IB4)-binding subpopulation of nociceptive primary afferents led to changes in NK-1r expression by the different lamina I cell types. We injected IB4 conjugated to saporin (SAP) into the left sciatic nerve of anesthetized male Sprague Dawley rats to specifically lesion IB4-positive non-peptidergic nociceptive C-fibers. Cholera toxin subunit B (CTB) was injected into the parabrachial nucleus to label lamina I projection neurons. Animals were tested for thermal and mechanical sensitivity and sacrificed from 2 weeks to 2 months post-lesion. We cut horizontal sections of spinal segments L4 and L5 and processed the tissue for IB4 binding and NK-1r and CTB immunoreactivities using immunofluorescence. IB4-SAP treated animals showed no behavioral changes compared to sham animals when tested for thermal (Hargreaves test), mechanical allodynia (von Frey test) or mechanical hyperalgesia (pin prick test) at any of the time points studied. Compared to the contralateral side and the sham group, lamina I projection neurons in the IB4-SAP treated group revealed an ipsilateral increase in the expression of NK-1r by the fusiform and multipolar neuronal populations. Nonetheless, there was no significant change in the percentage of pyramidal neurons which expressed NK-1r, which remained very low on the ipsilateral side of the IB4-SAP treated group. From these results, we infer that a loss of non-peptidergic afferents does not induce a phenotypic switch in the pyramidal neurons. However, the increase in NK-1r immunoreactivity in lamina I fusiform and multipolar neurons suggests that these cell populations may be important in maintaining the nociceptive responses in the absence of the IB4-positive non-peptidergic afferents. Finally, we suggest that a chronic pain state may be required for the de novo expression of NK-1r by pyramidal neurons.

Related Products: IB4-SAP (Cat. #IT-10)

Kozicz T, Xu L, Geenen B, Gaszner B, Kovacs K, Roubos E (2011) Leptin-receptor-expressing neurons in the non-preganglionic Edinger-Westphal nucleus regulate white and brown adipose tissue. Neuroscience 2011 Abstracts 822.19. Society for Neuroscience, Washington, DC.

Summary: Leptin, produced by white adipose tissue (WAT), is a key factor that regulates food intake and energy expenditure in vertebrates. It conveys information about fat storage in the periphery to the brain. The leptin receptor long form (LepRb) can be found in the non-preganglionic Edinger-Westphal nucleus (npEW) in the midbrain, which is the main site of urocortin 1 (Ucn1) production in the brain. In both mice and rats, intraperitoneal administration of leptin induces an increase in Ucn1 expression in the npEW whereas in mice that lack LepRb (db/db mice), the npEW contains considerably reduced amount of Ucn1. The npEW also responds to acute thermal exposure, indicating a role of this nucleus in thermoregulation. Brown adipose tissue (BAT) is critical to maintain homoeothermia and is centrally controlled via sympathetic outputs. A recent study demonstrates a projection from EW to BAT by using retrograde tracer pseudorabies virus (PRV). In our study, using PRV injection into the WAT of rats, we identified PRV-labeled Ucn1 neurons in the npEW, indicating a connection from npEW to WAT. In order to analyze the involvement of the npEW in the regulation of sympathetic WAT and BAT outputs, we performed the experiment using the neurotoxin saporin. When conjugated to leptin (Lep-SAP), Lep-SAP can selectively kill LepRb-expressing neurons. Wister rats were given injection of either Lep-SAP or a control blank saporin (B-SAP) into the left npEW Results showed that injection of Lep-SAP significantly blunted Ucn1 expression in the npEW. The weights of WAT and BAT were analyzed on both sides. The WAT and BAT weights were increased significantly on the contralateral side in Lep-SAP compared with B-SAP injected rats, however not different on the ipsilateral side. Interestingly, we observed that both WAT and BAT weighed more on the ipsilateral than the contralateral side only in the B-SAP animals. We will further test the effect of lesioning npEW neurons on the function of WAT and BAT by assessing specific WAT and BAT markers by RT-PCR and histology. Taken these data together, we provide evidence that LepRb-expressing neurons in the npEW regulate BAT and WAT most probably via sympathetic circuits.

Related Products: Leptin-SAP (Cat. #IT-47), Blank-SAP (Cat. #IT-21)

Jeong D, Chang W, Lee D, Chang J (2011) Decrease of GABAergic markers and Arc protein expression in the frontal cortex by injection of intraventricular 192 IgG-saporin. Neuroscience 2011 Abstracts 878.08. Society for Neuroscience, Washington, DC.

Summary: Previous studies used 192 IgG-saporin to study cholinergic function because of its facility for selective lesioning; however, results varied due to differences in the methods of administration and behavioral tests used. We investigated whether intraventricular injections of 192 IgG-saporin were suitable to make a dementia animal model for the evaluation of therapeutic drugs or electrical stimulation techniques. We examined the effects of 192 IgG-saporin using the Morris water maze, immunochemistry, and western blotting. Animals were examined 2 weeks after intraventricular injection of 192 IgG-saporin (0.63 µg/µl, 6 µl, 8 µl, and 10 µl) or phosphate buffered saline (8 µl). In the acquisition phase of the Morris water maze, the latencies of the injection groups were significantly delayed, but recovered within 1 week. In the probe test, two of four indices (time in the platform zone and the number of crossings) were significantly different between the control group and the group injected with 8 µl of 192 IgG-saporin. Immunohistochemistry revealed the extent of cholinergic destruction that was apparent in the basal forebrain of all 192 IgG-saporin injected rats. We found significantly decreased activity-regulated cytoskeleton associated protein (Arc) and glutamate decarboxylase (GAD) expression in the frontal cortex (8 µl and 10 µl groups), but not in the hippocampus, using western blotting. Further, spatial memory impairment was associated with cholinergic basal forebrain injury as well as fronto-cortical GABAergic hypofunction and synaptic plasticity deceleration. We conclude that intraventricular injection of 192 IgG-saporin is a suitable method for making a rat model of dementia.

Related Products: 192-IgG-SAP (Cat. #IT-01)

Lee J, Jeong D, Chang J (2011) The effects of basal forebrain cholinergic neuron on novel object recognition. Neuroscience 2011 Abstracts 878.10. Society for Neuroscience, Washington, DC.

Summary: Medial septum and basal nucleus areas of the basal forebrain project cholinergic neurons to the frontal cortex and the Hippocampus.And degeneration of the cholinergic basal forebrain neurons is a common feature of Alzheimer’s disease (AD) has been correlated with cognitive decline. This research was studied to verify the effects of cholinergic neuron in basal forebrain to the role of the hippocampus and the frontal cortex on recognition through recognition test and immunohistochemistry after damaging cholinergic neuron of the basal forebrain by intraventricular injection of 192 IgG-saporin. 192 IgG-saporin of 8ul (0.63ug/ul) was injected to the bilateral lateral ventricle of rats. After 2 weeks, novel object recognition (NOR) test was conducted to elucidate damage of cholinergic neuron. In the NOR test, rats are exposed to two identical objects for 15 minutes in empty plastic box (60cmx60cmx30cm). After 3 hours, they are reintroduced to the same object and a new novel object for 10 minutes. This procedure was repeated for 4 days After completing the behavioral experiment, the ChAT of cholinergic neuron in the basal forebrain was ascertained to confirm with immunohistochemistry if cholinergic neuron was damaged. In NOR test, the lesion group with 192 IgG-saporin showed 10% lower novel object preference than normal group. However, this rate is not that significant value enough to elucidate behavioral difference between normal group and lesion group. In immunohistochemistry, the number of cholinergic neuron was remarkably decreased in basal forebrain. According to both of the NOR test and Immunohistochemistry in the condition under lesion, Cholingergic input to hippocampus and frontal cortex from basal forebrain affects recognition somewhat, however the effect is not so essential.

Related Products: 192-IgG-SAP (Cat. #IT-01)

Bhide NS, Dickinson S, Feinberg E, Mohamed M, Dupre K, Eskow-Jaunarajs K, Lindenbach D, Ostock C, Bishop C (2011) Norepinephrine denervation by dopamine beta-hydroxylase saporin impacts L-DOPA efficacy and side effects in a hemi-parkinsonian rat model. Neuroscience 2011 Abstracts 883.20. Society for Neuroscience, Washington, DC.

Summary: Dopaminergic neurodegeneration in Parkinson’s disease (PD) is accompanied by concomitant loss in the norepinephrine (NE) system. The exact contribution of NE denervation in the development of PD remains elusive. Recently, we demonstrated that NE neurons may contribute to the efficacy and side effects of L-DOPA, however, to better mimic NE loss observed in PD we employed the selective NE neurotoxin dopamine beta hydroxylase saporin (DHB saporin) and evaluated its effects on the anti-parkinsonian efficacy of L-DOPA and the development & expression of L-DOPA induced dyskinesia (LID). To do so, hemiparkinsonian adult Sprague-Dawley rats were exposed to intraventricular injections of either vehicle or DHB saporin. Three weeks later, animals were primed with L-DOPA (4mg/kg) for days 1-7 and L-DOPA (12 mg/kg) for days 9-15. During this period animals were monitored for motor-performance, a marker for L-DOPA’s anti-parkinsonian efficacy, and dyskinesia measured using Abnormal Involuntary Movements (AIMs) scale. Further, sensitivity of primed animals to different doses of L-DOPA (ranging from 2 to 12 mg/kg) was assessed. Results indicate that NE denervation resulted in reduced anti-parkinsonian efficacy of L-DOPA, but not the development of dyskinesia. In fully primed rats, NE denervation attenuated dyskinetic responses to L-DOPA when compared to animals with an intact NE innervation. These findings suggest that the NE system significantly modulates the anti-parkinsonian effects of L-DOPA and the expression of LID and indicate the importance of understanding the mechanisms by which NE modifies basal ganglia function in PD.

Related Products: Anti-DBH-SAP (Cat. #IT-03)

Lee C, Rajkumar R, Suri S, Chin WM, Dawe GS (2011) The nucleus incertus contributes to the anxiety-like behaviour in rats. Neuroscience 2011 Abstracts 901.09. Society for Neuroscience, Washington, DC.

Summary: The nucleus incertus (NI), the principal source of relaxin-3 (Rln3) in the brain, is found in the periventricular gray, ventral and medial to the posterodorsal tegmental nucleus (PDTg). Several neuroanatomical studies have indicated that the NI projects to putative correlates of anxiety, especially the amygdala. Relaxin family peptide receptor type-3 (Rxfp3), the native receptor for Rln3, is expressed in the amygdala. These studies have hence predicted that the NI is strategically located to control neural circuits that underlie anxiety-like behaviour in rodents. Presence of Rln3-immunoreactive nerve fibres in the amygdala suggested the involvement of the Rln3/Rxfp3 system. Corticotrophin-releasing factor receptors type-1 (Crfr-1), one of the important anxiolytic drug targets, are prominently expressed in the NI neurons. Based on the aforesaid anatomical and receptor distribution reports, the present investigation was designed to clarify the function of the NI in anxiety-like behaviour of rats. We hypothesized that lesioning of the NI and the resulting decrease in Rln3 would affect the regulation of stress and anxiety response in rats. Firstly, the effect of NI neuron ablation, by CRF-Saporin toxin, on fear conditioning and elevated plus maze (EPM) exploration paradigms was evaluated. Secondly, the firing rates of NI neurons as the rat explored the EPM were assessed. Lastly, the effects of high frequency simulation of the NI on the expression of immediate early genes (IEG) in the amygdala were studied. The results revealed that, in a cued fear conditioning paradigm, NI-lesioned rats exhibited greater fear, indicated by longer freezing periods in the test phase, than sham-lesioned rats. Likewise, in the EPM, NI-lesioned rats made fewer entries into and spent less time in the open arms demonstrating an anxious phenotype. In addition, the NI also showed distinct firing patterns in the open and closed arms of the EPM. Stimulation of the NI activated the medial amygdaloid (MeA) nucleus as indicated by the increased expression of markers of neuronal activation. To sum up, the present study shows a significant contribution of the NI and NI-MeA pathway in the anxiety-like behaviour of rats. It also suggests that the NI and/or Rln3 have a role in the regulation of anxiety-like behaviour, implicating them as targets for anxiety-related disorders.

Related Products: CRF-SAP (Cat. #IT-13)

Roach EB, Hussain Shuler MG (2011) Cholinergic reinforcement and temporal learning in rodent visual cortex. Neuroscience 2011 Abstracts 608.16. Society for Neuroscience, Washington, DC.

Summary: The idea that neuromodulators act as reinforcement signals has an intricate scientific history, including the well-characterized analogue of prediction error relayed by midbrain dopamine neurons. Neuromodulators released from discrete nuclei are poised to broadcast to many brain regions at once, and so it is an appealing concept to investigate other neuromodulatory systems within a reinforcement learning framework. Reward timing activity, a neural reflection of operantly learned stimulus-reward intervals in the primary visual cortex (V1), offers a tractable in vivo model to examine the role of candidate neuromodulators in temporal reward learning. Reward timing was first characterized in rats trained to lick a delivery tube to receive water rewards, where stimulation to one eye indicated reward availability after x licks, while stimulation to the other eye required y licks. Simultaneously recorded activity in V1 indicated that single unit responses evolve from reporting only visual characteristics to showing persistent increased/decreased firing or peak activity corresponding to the time of anticipated reward. Individual neurons report one interval or the other, even those with binocular peri-stimulus responses, arguing that reward timing is learned locally within V1. Theoretical work suggests that the local expression of reward associated intervals requires an interaction between the visually-evoked network response and a reinforcement signal conveying the time of reward. Based on anatomical and neurophysiological evidence, we hypothesized that cholinergic input from the basal forebrain (BF) could provide such a reward signal to V1. To test its necessity, BF cholinergic innervation in V1 was lesioned — using the selective neurotoxin 192 IgG-saporin — prior to changing the experimental policy between cues and associated reward delays. This allowed an examination of two potential roles for BF cholinergic input: in expressing previously learned intervals and in acquiring information about new intervals. We found that neurons from saline-infused controls, but not lesioned animals, shifted as a population to report the new, behaviorally relevant intervals (Kolmogorov-Smirnov, p < 0.05). Importantly, neurons from lesioned animals continued to report the previously learned intervals, suggesting that BF cholinergic input is required to learn, but not express, reward timing. These results support the notion that acetylcholine released from BF afferents acts as a reinforcement signal that guides cortical network plasticity.

Related Products: 192-IgG-SAP (Cat. #IT-01)

Paolone G, Lamy D, Sarter M, Lee T (2011) Cognitive performance-associated increases in cholinergic neurotransmission also serve as a circadian signal to sustain performance-induced diurnal activity patterns. Neuroscience 2011 Abstracts 610.12. Society for Neuroscience, Washington, DC.

Summary: Daily practice of a sustained attention task (SAT) during the light phase of the light/dark cycle causes a stable, entrained, diurnal behavioral activity pattern (Gritton et al. 2009). As SAT performance is mediated by increases in cortical cholinergic neurotransmission, this experiment assessed levels of acetylcholine (ACh) release across the light and dark cycle of animals that previously performed the SAT at a fixed time. Circadian behavioral activity was recorded, and prefrontal ACh release was measured, using microdialysis, beginning on the third day following the last SAT session. SAT practice took place in either the light phase [ZT4], the dark phase [ZT16], or in a constant light condition [LL]. A control group practiced a daily fixed interval [FI-9] schedule of reinforcement at ZT4. A second control group was handled at randomly selected times but was neither water-deprived nor performed a task [NP]. Dialysates were collected, in a new environment, for 180 min total, beginning 90 min before the onset of prior task practice and again during the equivalent time period twelve hours later. For all animals, ACh release levels were higher during the dark phase. In SAT-performing animals, ACh levels increased for 45 min at ZT4 and ZT16. In addition, the ZT4 animals’ behavioral activity was robustly increased during this interval. Animals trained at ZT 4 reversed back to a nocturnal activity pattern 8-10 days after cessation of SAT practice, coinciding with the loss of the task time-synchronized cholinergic activity. In order to determine the necessity of these prior task period-synchronized release events for maintaining diurnal activity patterns, basal forerbain cholinergic neruons were lesioned by intra-basalis infusion of 192 IgG-saporin. As was expected, this lesion impaired SAT performance. Furthermore, following cessation of daily SAT practice, prior performance-period synchronized cholinergic release events were abolished in lesioned animals. Moreover, the lesion triggered a rapid post-performance return to a nocturnal acitvity pattern. Collectively, these results indicate that SAT performance-associated increases in prefrontal cholinergic activity not only support SAT performance but also contribute to cognition-induced diurnality. Furthermore, circadian control of cholinergic activation optimizes task performance as well as the generation of a cholinergic zeitgeber signal. In conclusions, the brain’s clocks and increases in cortical cholinergic neurotransmission interact bidirectionally to sustain cognitive performance and performance-evoked diurnal activity patterns.

Related Products: 192-IgG-SAP (Cat. #IT-01)