sfn2009

Freiria-Oliveira AH, Blanch GT, De Paula PM, Colombari E, Menani JV, Colombari DS (2009) Role of A2 noradrenergic neurons and angiotensinergic mechanisms on hypotension induced by hemorrhage. Neuroscience 2009 Abstracts 467.18/DD70. Society for Neuroscience, Chicago, IL.

Summary: The A2 catecholaminergic neurons in the commissural subdivision of the nucleus tractus solitarii (cNTS) are activated by hemorrhage. However, the role of these neurons on the cardiovascular adjustments to hemorrhage is not fully understood. In the present study we investigated the effects of A2 noradrenergic neuron lesion alone or combined with the blockade of angiotensinergic mechanisms on the recovery of blood pressure after hemorrhage. Male Holtzman rats (280-320 g) anesthetized with ketamine combined with xylazine were submitted to lesions of dopamine-beta-hydroxilase (DβH)-containing neurons in the cNTS achieved with injections of anti-DβH-saporin (12.6 ng/60 nl, n=6-8) or sham lesions (injection of immunoglobulin-G-saporin, 12.6 ng/60 nl, n=6). Changes in blood pressure to hemorrhage were tested 30 days after lesions. Immunohistochemistry for tyrosine-hydroxilase was performed to confirm the efficacy of DβH neuron lesion in the cNTS. Two days before tests, femoral artery and vein were cannulated under ketamine and xylazine anesthesia. Hemorrhage consisted in four blood withdrawals (2 ml/300 g body weight, every 10 min) in conscious rats. Immediately after the 4th blood withdrawal, the hypotension was similar in A2-lesioned and sham-lesioned rats (-62 ± 7 mmHg and -73±7 mmHg, respectively). However, A2-lesioned rats rapidly (20 min) recovered from hypotension (-7±2 mmHg), while sham rats did not completely recover from hypotension until the end of experiment (60 min after the 4th blood withdrawn, -20±3 mmHg). The pre-treatment with losartan (angiotensin type 1 receptor antagonist, 10 mg/kg of body weight, iv) impaired the recovery of blood pressure by A2-lesioned rats (-29 ± 4 mmHg and -28 ± 3 mmHg, 20 and 60 min after the 4th blood withdrawal). In sham rats, the treatment with losartan also reduced the partial recovery of blood pressure at the end of the test (-39±6 mmHg, vs. sham control: -20±3mmHg), however, losartan did not affect the hypotension 20 min after the 4th blood withdrawal (-30± 6 mmHg vs. sham control: -35 ± 9 mmHg). The results suggest that A2 noradrenergic neuron lesion in the cNTS facilitates the recovery of hypotension after hemorrhage, probably increasing the action of angiotensinergic mechanisms.

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

Macleod JE, Ackerman CM, Bucci DJ (2009) Contributions of the medial prefrontal cortex to negative occasion setting. Neuroscience 2009 Abstracts 477.2/FF61. Society for Neuroscience, Chicago, IL.

Summary: The medial prefrontal cortex of rats has a role in many aspects of cognitive function, and especially forms of inhibitory learning. Recent research has revealed heterogeneous functions of the prelimbic (PL) and infralimbic (IL) regions of the medial prefrontal cortex in modulating response inhibition. In a recent study, we tested the effects of separate neurotoxic lesions of the PL or IL in a serial feature negative discrimination paradigm (negative occasion setting). Rats received daily training sessions consisting of 16 trials: on 4 trials in each session, a tone was presented and followed by food reward; on the remaining trials, the tone was preceded by a visual stimulus and not reinforced. Our results indicate that PL but not IL is necessary for learning the discrimination. A second study was conducted to investigate the effects of these lesions on rats that were first extensively trained in this task. We found that rats that had been trained for 30 days prior to receiving PL or IL lesions were still able to perform the task as well as controls. Therefore, PL lesions disrupt acquisition but not performance of a serial feature negative discrimination. This same task has been used in our laboratory to investigate the effects of nicotine on learning. We have shown that nicotine-treated rats exhibit greater discrimination between the two trial types as evidenced by less frequent responding during non-reinforced trials, and learn the discrimination in fewer sessions than control rats. In addition, rats receiving nicotine showed an increase in rearing behavior during the presentation of the light, suggesting nicotine enhanced attention to the visual stimulus. One possible critical site of action for nicotine’s effects is the medial prefrontal cortex. Research in other laboratories utilizing other training procedures suggest that cholinergic activity in the medial prefrontal cortex is critical for attending to behaviorally relevant stimuli, and have implicated the rat PL in visual attention as well as inhibiting prepotent, goal oriented responses. We investigated the contribution of the cholinergic PL to learning the serial feature negative discrimination task by training rats that had received infusions of 192-IgG-saporin into PL to remove cholinergic input from the basal forebrain. No differences between control and lesion rats were observed. Taken together, the results suggest that PL is necessary for acquisition of a serial feature negative discrimination, although the basal forebrain cholinergic input into this region is not required to sufficiently learn the task.

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

Laplante FP, Dufresne M, Lappi DA, Sullivan RM (2009) Depletion of cholinergic neurons in the nucleus accumbens impairs dopamine function in the prefrontal cortex in the rat. Neuroscience 2009 Abstracts 341.7/O16. Society for Neuroscience, Chicago, IL.

Summary: Studies of post mortem schizophrenic brains have revealed a selective loss of cholinergic interneurons, most pronounced in the ventral striatal region. We have previously shown in the rat, that a novel saporin immunotoxin coupled with an antibody targeting choline acetyltransferase (ChAT) and microinjected (0.5 _g/_l; 0.5 _l) into the nucleus accumbens (N. Acc) of adult rats, reduces the number of cholinergic neurons in N. Acc. by 40-50 %. Such lesions result in a markedly heightened response to the locomotor activating effects of amphetamine and impair prepulse inhibition of the acoustic startle response. We proposed that this local cholinergic deficit leads to a hyperresponsiveness in subcortical dopamine (DA) systems of relevance to schizophrenic symptomatology. Presently, we hypothesize that the same local cholinergic defect may trigger broader changes in cortical/subcortical networks, specifically prefrontal cortex (PFC) deficits in DA-mediated functions, also proposed in schizophrenia. Young adult male Srpague-Dawley rats were injected bilaterally in the N. Acc. as described above with either the cholinergic immunotoxin or vehicle. Two weeks later, they were trained in a working memory task dependent on PFC function, using the delayed alternation paradigm in the T-maze. Lesioned rats took significantly longer to reach criterion performance during training than controls. During testing, lesioned rats were significantly impaired in the percentage of correct arm choices across delay intervals, but especially with longer (40 sec) delays. The same animals were then implanted with voltammetric recording electrodes in the ventromedial PFC to examine the increases in in vivo extracellular DA release in response to a brief tail pinch stress. Lesioned rats showed a significantly reduced activation of the mesocortical DA system compared to controls. Taken together, the data suggest that reduction in the density of cholinergic neurons in the N. Acc also triggers deficits in prefontally-mediated function known to be under mesocortical DAergic regulation. This raises the possibility that ventral striatal cholinergic deficits may be causally linked to cortical/subcortical functional imbalances proposed to exist in schizophrenia.

Related Products: Anti-ChAT-SAP (Cat. #IT-42)

Lima T-Z, Blanco MM, Bueno MA, Dos Santos Junior JG, Bargieri DY, Mello LE (2009) The influence of cholinergic degeneration on the progression of Alzheimer’s disease and its action in determining the outcome of lithium treatment. Neuroscience 2009 Abstracts 139.26/D36. Society for Neuroscience, Chicago, IL.

Summary: A substantial loss of cholinergic innervation in the hippocampus and cerebral cortex is universally accepted as a typical feature of Alzheimer’s disease (AD). Cholinergic deafferentation is an often, but not a constant phenomenon in AD and its contribution to the progression of disease is not completely understood. The present work was aimed to evaluate the effect of cholinergic deafferentation on cognitive decline and on Amyloid-b (A_) metabolism and how this outcome is modulated by lithium. To this end rats were subjected to neonatal intracerebroventricular injection of 192 IgG-saporin (an immunotoxin selective to cholinergic neurons). Three months after surgery animals were evaluated in Morris Water Maze (MWM) and then entered a three months long lithium (LiCl) or control treatment. At the end of treatment, animals were once again tested in MWM and their cerebral cortical A_ levels were measured. We found that working memory impairment arises earlier and it is also more severe than reference memory related deficits. The cognitive performance was only slightly influenced by LiCl treatment. Furthermore we found that the cholinergic denervation produced by neonatal IgG-sap infusion did not modify A_ levels or its aggregation state. Moreover lithium increased the levels of A_1-42 despite decreasing the amount of A_1-40, an effect that is dependent of cholinergic integrity. These data suggest that the contribution of cholinergic deafferentation, which occurs over the progression of disease, to the amyloigenesis is likely to be modest in AD brain. In addition the effects of lithium treatment presented here imply in avoiding its use as prophylactic propose for AD and in AD cases without marked cholinergic degeneration.

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

Wiley RG, Kline IV RH, Lemons LL (2009) Role of galanin receptor-expressing dorsal horn neurons in nocifensive reflex responses to heat. Neuroscience 2009 Abstracts 170.17/X19. Society for Neuroscience, Chicago, IL.

Summary: Spinal intrathecal (i.t.) galanin has been reported to be antinociceptive in some situations. Using lumbar i.t injections of galanin, coupled to the ribosomal inactivating peptide, saporin, to selectively destroy spinal dorsal horn cells that express galanin receptors, we sought to determine the role of galanin receptor-expressing dorsal horn neurons in reflex nocifensive hotplate behavior. Rats were injected into lumbar CSF with either 500 ng or 750 ng Gal-sap or saline, then tested over several weeks on the hotplate at 44o, 47o and 52oC. Gal-sap increased hindpaw withdrawal latencies only to 44oC and decreased the amount of responding on both 44o and 47oC hotplates. Morphine (5 mg/kg, s.c.) twenty minutes before 44°C hotplate testing slightly increased initial response latency and significantly decreased responding of the control rats. The antinociceptive effect of morphine in the Gal-sap rats was approximately additive with the antinociceptive effect of Gal-sap. Mustard oil applied to the dorsal hindpaws significantly increased responding on the 44°C hotplate in control rats, but produced less of an increase in Gal-sap rats. Topical capsaicin to hindpaw plantar skin reduced control, but not Gal-sap, responses on the 44°C hotplate. These results suggest a role for galanin receptor-expressing dorsal horn neurons in modulation of nociception that is unique, different from several other types of dorsal horn neurons and suggests a strategy for augmenting opiate drug effect.

Related Products: Galanin-SAP (Cat. #IT-34)

Lemons LL, Wiley RG (2009) Role of galanin receptor-expressing dorsal horn neurons in operant nocifensive responses. Neuroscience 2009 Abstracts 170.18/X20. Society for Neuroscience, Chicago, IL.

Summary: Selective destruction of galanin receptor-expressing dorsal horn neurons using the targeted cytotoxin, galanin-saporin (gal-sap), reduced reflex nocifensive hotplate responses, particularly at 44° C (see adjacent poster). The antinociceptive effect of gal-sap was additive with morphine (5 mg/kg, s.c.) in reducing hotplate responses. While these findings are provocative, inferences about analgesia also require information on cerebral processing of nociceptive information, such as obtained from operant nocifensive responses. We therefore sought to determine the effects of lumbar intrathecal gal-sap on nocifensive operant responses. Thirteen Long Evans female rats were injected with either 500 ng gal-sap or 500 ng blank-sap and tested on the escape test at several temperatures. The escape task consists of a two-chambered box; one side is dark with a thermal floor while the other side is brightly lit with a room temperature shelf. Gal-sap treated rats escaped from the thermal plate to the escape shelf less than controls. The difference was particularly striking at 25°, 38°, 44°, 45°, and 47°C. Morphine effects on escape responses was tested at 44°C. Thirty minutes before testing, rats were injected subcutaneously with either 0, 0.5, 1.5 or 4.5 mg/kg morphine. The testing was done over four days such that every rat was tested at each dosage in a counterbalanced fashion. 1.5mg/kg of morphine significantly reduced the escape duration of the blank-sap control rats, but not the gal-sap rats. The 4.5mg/kg dosage completely eliminated escape responding in both control and gal-sap rats. In summary, Gal-sap rats showed reduced nocifensive reflex responding on the hotplate tests, which was further reduced in essentially additive fashion by 5mg/kg of morphine. The Gal-sap treated rats also showed reduced escape behaviors at 44°C in the operant escape test, but were less sensitive to 1.5 mg/kg of morphine than the control rats. These results differ from the effects of NPY-saporin and dermorphin-saporin, cytotoxins also targeted at dorsal horn interneurons, suggesting that selective destruction of galanin receptor-expressing superficial dorsal horn neurons is analgesic and that galanin-receptor-expressing dorsal horn interneurons play a unique role in nociceptive processing.

Related Products: Galanin-SAP (Cat. #IT-34)

Jaime S, Perez Cordova MG, Hernandez S, Colom L (2009) Immunolesions of medial septal GABAergic neurons. Neuroscience 2009 Abstracts 241.8/I15. Society for Neuroscience, Chicago, IL.

Summary: Epilepsy is a neurodegenerative condition characterized by spontaneous recurrent seizures that are triggered by excessive electrical activity due to changes in neurological functions. One of the most common forms of epilepsy is Temporal Lobe Epilepsy (TLE) in which seizures originate in limbic structures as hippocampal and/or para-hippocampal areas. Principal cell (i.e. pyramidal cells) activity is indirectly regulated by rhythmic inputs from GABAergic neurons in the septal region of the basal forebrain which selectively innervate inhibitory hippocampal interneurons. In previous studies, using the pilocarpine model of TLE, we have demonstrated that the septum plays an antiepileptic role and that medial septum GABAergic neurons degenerate in the epilepsy process. Thus, damage of medial septum GABAergic neurons may contribute to epileptogenesis. The purpose of this study is to investigate the role of medial septum GABAergic neurons in excitability control and epileptic activity generation. For this purpose, anti-GAT1-SAP (3µL at 325ng/µL) was stereotaxically injected in the medial septum of Sprague Dawley male rats to selectively destroy this neuronal population and investigate the subsequent functional changes. Analysis was performed using stereological approaches which revealed a significant reduction in cell count between treated (anti-GAT1-SAP) and saline-injected control rats (8591.38±941.65 and 25609.87±407.73 respectively; (Student’s t-test; p<0.05). In conclusion, our preliminary results show that the single injections of anti-GAT1-SAP selectively lesions most of the medial septum GABAergic neurons, providing a powerful tool to study the role of these neurons in the control of hyperexcitability states. Studies underway involve the investigation of the functional alterations produced by the selective destruction of MS GABAergic neurons.

Related Products: GAT1-SAP (Cat. #IT-32)

Datta S, Chatterjee K, Kline IV RH, Wiley RG (2009) CCK receptor- expressing dorsal horn neurons: Role in pain and morphine analgesia. Neuroscience 2009 Abstracts 265.13/Z37. Society for Neuroscience, Chicago, IL.

Summary: Spinal intrathecal cholecystokinin (CCK) has anti-opiate activity, and the CCK antagonist, proglumide potentiates opiate analgesia. In the present study, we sought to determine the effects of selectively destroying CCK receptor-expressing lumbar dorsal horn neurons using the targeted cytotoxin, CCK-saporin on reflex and operant nocifensive responses to heat, and on the actions of systemic morphine and naloxone. Exp. 1: Adult, female rats were injected into the lumbar CSF with either 1500 ng of CCK-sap (n=7) or blank (control nonsense peptide)-saporin (n=6). Exp. 2: rats were pre-injected intrathecally with 1 ug of proglumide (CCK antagonist) followed by 1500 ng CCK-sap (n=4) or only CCK-sap (1500 ng; n=4). Rats were then tested on the hotplate at 44°C and 47°C and on an operant thermal preference task (TPT) using a shuttle box where the floor on one side was 15°C and the other 45°C. Morphine was tested in the TPT using 0, 0.5, 1.5 and 2.5 mg/kg s.c. 4-8 weeks post-toxin. Naloxone (0 vs 0.8 mg/kg s.c) was also tested in the TPT. In Exp. 1, the CCK- sap group showed decreased hotplate reflex responses, but decreased time on the 45°C side in the TPT. In Exp. 2, CCK-sap only rats also showed greater heat aversion in the TPT. In both Exps, CCK-sap groups demonstrated greater heat aversion (less analgesia) than either control group after morphine in the TPT. After naloxone, both control groups, but not the CCK-sap rats, showed increased heat aversion (hyperalgesia). We interpret these results as showing that selective destruction of CCK receptor- expressing superficial dorsal horn neurons increases nocifensive reflex responses to aversive heat and produces thermal hyperalgesia while decreasing the effects of both morphine and naloxone suggesting a complex role for CCK receptor-expressing dorsal horn neurons in modulation of nociception and opiate drug action.

Related Products: CCK-SAP (Cat. #IT-31)

Vetrivelan R, Anaclet C, Fuller PM, Yoshida K, Lu J, Saper CB (2009) Comparison of sleep-wake changes after lesions of two sleep-promoting cell groups in the preoptic region in rats. Neuroscience 2009 Abstracts 277.2/EE14. Society for Neuroscience, Chicago, IL.

Summary: Previous studies have shown that two cell groups within the preoptic region viz., median preoptic nucleus (MnPO) and ventrolateral preoptic nucleus (VLPO) show c-Fos expression during spontaneous sleep. However, while lesions of the VLPO have been shown to cause sleep loss, the effects of MnPO lesions on sleep have not been available to date. We therefore performed cell-body specific lesions of these two nuclei using the toxin orexin-saporin and studied the spontaneous sleep-wake behavior in rats. We found that the animals with more than 70% cell loss in the VLPO (n=15) showed a 31% increase in wakefulness (61.03±1.15% in VLPO-lesioned animals vs 46.53±0.55% in controls, P <0.001) and a concomitant reduction in non rapid eye movement (NREM) sleep and REM sleep. There was also a significant reduction in the average NREM sleep episode duration (120±6.57 Sec Vs 143.04 ± 4.53 sec in control animals, P < 0.01) in rats with VLPO lesions. On the other hand, lesions involving 80-90% cell loss in the MnPO (n=6) produced a moderate 15% increase in wakefulness (53.8±1.09% vs 46.53±0.55% in controls. P<0.001). Although the NREM sleep episode duration was reduced in these animals (126 ± 6.61 Sec vs 143.04 ± 4.53 sec in control animals, P = 0.06), it did not reach statistical significance. The extent of the lesions in the present study was estimated by an individual blind to the experimental conditions and the sleep results. Although specific cell groups (MnPO or VLPO) were carefully targeted, partial damage (10-20%) to the other cell group was often encountered. Nevertheless, our results clearly demonstrate that while the MnPO plays an important role in the regulation of sleep, the VLPO plays a substantially greater role.

Related Products: Orexin-B-SAP (Cat. #IT-20)

Morin LP, Studholme KM (2009) Saporin lesions that target suprachiasmatic cells bearing NPY receptors eliminate or greatly impair circadian rhythm generation and entrainment. Neuroscience 2009 Abstracts 278.7/EE49. Society for Neuroscience, Chicago, IL.

Summary: General destruction of the SCN caused by electrical lesions produce loss of circadian rhythmicity and entrainment. More specific, cell-directed lesion methods, such as the use of NMDA as a neurotoxin, have not been successful. Here, we describe the use of the ribotoxin, Saporin (SAP), to kill specific types of SCN neurons and show the effects of such selective lesions on the hamster circadian locomotor rhythm. Adult male golden hamsters were injected bilaterally with 200 nL of a SAP/neuropeptide conjugate into the SCN bilaterally. The neuropeptides were neuropeptide Y (NPY), cholecystokinin (CCK) or substance P (SP). NPY terminals are distributed throughout the SCN; CCK and SP cells are present in the SCN and there have been NPY and SP receptors described in the hamster SCN. SAP/NPY (N=10) treatment caused arrhythmicity in 4 animals under LD conditions and 4 others became arrhythmic when transferred to DD. Arrhythmicity occurred in 1/9, 0/8 and 0/10 animals treated with SAP/CCK, SAP/SP or vehicle. There was also a significant effect of treatment on the level of variability of the activity records as indicated by precision of activity onset (p<.008) and approximate entropy analysis of disorder within the running record (p<.004). The SAP/NPY group accounted for nearly all the between-group variability. The histology showed a large decrease in the number of SCN cells, but there were many cells remaining after SAP/NPY treatment. Care was taken to determine that the remaining cells were, in fact, neurons. Also, the brains of lesioned animals retained reasonably intact RHT, GHT and 5HT input pathways. Normal histology evaluated for NeuN, a neuronal antigen, showed that an unexpected pattern of NeuN-IR cells in the SCN of normal animals, with the majority of such neurons found in an area that includes the SCNce and the region dorsolateral. NeuN was heavily co-localized with calbindin-IR in cells of the SCNce, but not with VP- or VIP-IR. This distribution of SCN cells containing NeuN-IR was approximately the same in both mice and hamsters. Conclusions: (1) SAP/NPY lesions many, but not all SCN neurons; (2) Such lesions result in massive degradation of circadian rhythmicity; (3) The three main SCN input pathways remain essentially intact after SAP treatment; (4) NeuN-IR neurons are distributed in a novel pattern in the SCN of both mouse and hamster; (5) Presently unidentified SCN cells bearing NPY receptors are likely to be critical to the generation of cohesive circadian rhythms, whereas those bearing SP or CCK receptors are minimally, if at all involved.

Related Products: CCK-SAP (Cat. #IT-31), SSP-SAP (Cat. #IT-11), NPY-SAP (Cat. #IT-28)