References

Kolasa K, Harrell LE (2003) Combined lesions of central cholinergic and noradrenergic denervation in the rat using 192 IgG-saporin and DSP-4 as an animal model of Alzheimer’s disease. Neuroscience 2003 Abstracts 842.6. Society for Neuroscience, New Orleans, LA.

Summary: To better model the consequences of persistent cholinergic hypofunction observed in Alzheimer’s disease, medial septum (MS) lesions were made by using specific cholinotoxin 192-IgG saporin. In this study the effect of simultaneous hippocampal cholinergic denervation, induced by intraseptal injection of 192-IgG saporin, and central noradrenergic denervation, induced by systemic injection of DSP-4 (N-[2-chloroethyl]-N-ethyl-2-bromobenzylamine) was examined in the rat dorsal hippocampus. DSP-4, an adrenergic neurotoxin selective for locus coeruleus innervated brain regions, induced a decrease in norepinephrine (NE) concentration in hippocampus. MS lesions resulted not only in selective cholinergic denervation of hippocampus (CD; superior cervical ganglion removed to prevent ingrowth of peripheral NE fibers), but also ingrowth of NE fibers into the hippocampus (HI; superior cervical ganglion left intact). MS lesions also resulted in a significant loss of choline-acetyltransferase activity in HI and CD groups, and an increase in NE in the HI group. In the HI group, but not in CD or control groups, visualization of hippocampus revealed a dense NE innervation with fine NE fibers with varicosities. Combination of MS lesion and DSP-4 treatment resulted in a reduction of NE concentration in HI group, with concomitant decrease in visualization of NE fibers. Those that remained were thick with sparse varicosities, possibly derived from peripheral sympathetic ingrowth. Elevated NE concentration and NE fiber number following specific cholinergic lesions might reflect compensatory sprouting of both central and peripheral adrenergic fibers into the hippocampus. Thus, noradrenergic sprouting in response to cholinergic denervation of hippocampus might be a valuable model for studying mechanisms as well as the consequences of neuronal plasticity in the mature CNS.

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Hummer TA, Johnson AD, Givens B (2003) The effects of ethanol and cholinergic lesions on anxiety in mice. Neuroscience 2003 Abstracts 859.16. Society for Neuroscience, New Orleans, LA.

Summary: Ethanol’s anxiolytic properties in mice are well-documented as measured with an elevated plus-maze. The effect of IP ethanol injections and cholinergic lesions on anxiety was investigated. Animals were injected with 0.5 µg of the cholinergic immunotoxin anti-murine-p75-saporin or control saline into the right lateral ventricle. After recovery (14-17 d), mice were given an IP 1.6 g/kg ethanol (10% v/v) or vehicle injection. After 60 min, animals were placed on the elevated plus-maze for a 10 min session and then perfused. Mice displayed increased open arm entries and time following ethanol. Saporin did not effect measures of anxiety nor interact with the effects of ethanol. Choline acetyltransferase staining was performed to confirm the extent of the cholinergic lesioning, and ethanol assays confirmed the presence or absence of ethanol in the blood stream. The research demonstrates that, as measured in the plus-maze, the basal forebrain cholinergic system has no significant involvement in anxiety nor in ethanol’s anxiolytic effects.

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Farhadi HF, Lepage P, Forghani R, Friedman HC, Orfali W, Jasmin L, Miller W, Hudson TJ, Peterson AC (2003) A combinatorial network of evolutionarily conserved myelin basic protein regulatory sequences confers distinct glial-specific phenotypes. J Neurosci 23(32):10214-10223. doi: 10.1523/JNEUROSCI.23-32-10214.2003

Summary: The authors used intrathecal injections of 0.3 µg CTB-SAP (Cat. #IT-14) to induce spinal cord demyelination for the purpose of defining the regulatory network controlling myelin basic protein transcription in mice.

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Carstens EE, Iodi-Carstens M, Simons CT (2003) Reduced scratching in rats receiving intracisternal substance-saporin to ablate cervical superficial dorsal horn neurons that express NK-1 receptors. Neuroscience 2003 Abstracts 908.2. Society for Neuroscience, New Orleans, LA.

Summary: While glutamate and the neuropeptide substance P (SP) are involved in the spinal neurotransmission of nociceptive signals, little is known about transmitters involved in itch. We investigated a role for SP in itch by determining if scratching behavior is affected by selective neurotoxic destruction of cervical superficial dorsal neurons that express NK-1 receptors for SP. Sprague-Dawley rats received intracisternal microinjection of SP conjugated to saporin (SP-SAP; 2.27 μM/ 20 μl). Controls received saporin (SAP) only. At least 2 wk post-surgery, rats were tested for dose-related hindlimb scratching directed toward the site of intradermal microinjection of serotonin (5-HT; 50, 100 or 200 μg/10 μl) or saline (control) into the nape of the neck, with at least 1 wk between sessions for each dose. After the intradermal injection, rats were videotaped for 44 min. The numbers and durations of individual scratching bouts were counted and averaged for each dose. Rats receiving SAP exhibited a dose-related increase in scratching bouts similar to naïve rats. Rats receiving SP-SAP exhibited significantly reduced scratching (to ~38%) at all 5-HT doses. Individual bout durations (~2 s) did not vary significantly between groups or by dose of 5-HT. After behavioral testing, rats were perfused with fixative and caudal medullary and cervical spinal cord sections processed immunohistochemically for NK-1 receptors. Tissue from SAP-treated rats exhibited a normal distribution of pronounced NK-1 immunoreactivity in superficial layers of the dorsal horn at caudal medullary through C5 levels, while in SP-SAP-treated rats there was a complete absence of NK-1 immunoreactivity at these levels. These results indicate that SP plays an important role in neurotransmission from itch-signaling primary afferent fibers to second-order neurons in the superficial dorsal horn.

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Sarter MF, Martinez V, Bruno JP (2003) Lateralization of the attentional functions mediated via cortical cholinergic inputs. Neuroscience 2003 Abstracts 921.20. Society for Neuroscience, New Orleans, LA.

Summary: The role of basal forebrain (BF) corticopetal cholinergic projections in mediating attentional processing has been well established. For example, bilateral cortical cholinergic deafferentation produces robust impairments in attention. Neuropsychological and neuroimaging studies have postulated that attentional functions and capacities are mediated primarily via a lateralized, right-hemispheric network. The present study tested the general hypothesis that the attentional functions of cortical cholinergic inputs likewise are lateralized and thus that right-hemisphere cortical cholinergic deafferentation yields more severe attentional impairments. Rats were trained to perform an operant sustained attention task. Upon reaching criterion performance, unilateral cortical cholinergic deafferentation was produced by infusions of 192-IgG saporin into either the left or right BF. Compared with the performance of sham-operated animals and animals with left-hemispheric lesions, right-hemispheric cortical cholinergic deafferentation resulted in a persistent and selective decrease in the detection of signals (hits), mirroring the more potent but similarly selective effects of bilateral lesions. In contrast, left cortical cholinergic deafferentation did not affect hits but decreased the number of rejections in non-signal trials. These data extend previous studies suggesting that the integrity of the right cortical cholinergic input system is necessary for signal detection (Bushnell et al. 1998). Furthermore, the present data substantiate the assumption that the detection of signals and the rejection of non-signals are based on fundamentally different cognitive operations, and that the cholinergic mediation of these two operations is lateralized.

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Eichenbaum HB, Ross R, Raji A, McGaughy JA (2003) Noradrenergic, but not cholinergic, deafferentation of the infralimbic/prelimbic cortex impairs attentional set-shifting. Neuroscience 2003 Abstracts 940.7. Society for Neuroscience, New Orleans, LA.

Summary: Damage to the prefrontal cortex in humans produces deficits in the ability to shift attention to a previously irrelevant stimulus dimension (extra-dimensional shift; EDS) while sparing reversal and intra-dimensional shifts (a novel discrimination without a change in the relevant dimension; IDS). Data from human subjects has also shown the administration of noradrenergic agonists and antagonists disrupts EDS suggesting a role for coeruleal-cortical norepinephrine (NE). Usher and colleagues have proposed that high, tonic levels of NE may maximize behavioral flexibility to allow an optimal responsiveness to changes in the environment e.g. the predictability of reinforcement (Usher et al., Science 283, 1999). Based on these data, a loss of NE would be predicted to impair attentional set-shifting. In a rodent model of attentional set-shifting developed by Brown and colleagues, excitotoxic lesions of the infralimbic/prelimbic cortex (IL/PL) produced impairments in EDS but not IDS or reversal learning (J. Neurosci. 20, 2000). This study confirmed the importance of IL/PL to EDS, but did not address the role of NE in this type of cognition. In the current study, rats were infused with anti-dopamine beta-hydroxylase-saporin (NE-SAP)in IL/PL (0.01 µ g/ µl; 0.5 µl/hemisphere) to produce noradrenergic deafferentation, 192 IgG saporin (ACH-SAP; 0.01 µ g/ µl; 0.5 µl/hemisphere)to produce cholinergic deafferentation, or vehicle then tested in an attentional set-shifting task. NE-SAP rats were impaired in EDS but not in IDS or reversal learning. In contrast, ACH-SAP rats showed no impairment in any aspect of the task. The effect of DBH-SAP lesions on EDS support the hypothesis that NE, but not ACH, is critical to the adaptation of behavior to changes in reinforcement contingencies. The lack of effect of these lesions on reversal learning suggest the robustness of this effect may vary with the extent of behavioral adaptation required.

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Janssen WG, Andrews G, Tomey MI, Baxter MG, Morrison JH (2003) Combined bilateral perforant path lesions with lesions of the cholinergic system: an ultrastructural immunogold analysis of nmdar1 representation within the dentate gyrus. Neuroscience 2003 Abstracts 676.26. Society for Neuroscience, New Orleans, LA.

Summary: Alzheimer’s disease is characterized by deterioration of cholinergic input to the hippocampus, as well as degeneration of input from the entorhinal cortex to the dentate gyrus(DG). Studies have demonstrated an upregulation of the NMDA receptor subunit, NR1, following unilateral ablatement of the perforant path(pp). We hypothesized that cholinergic innervation might be essential for DG plasticity following pp ablation. Our study was designed to investigate the synaptic distribution of NR1 following combined 192 IgG-Saporin lesions of the medial septum/vertical diagonal band(MS/VDB) and bilateral(bilat) pp knife cut ablation. Animals received bilat-pp lesions 2-3 weeks days post MS/VDB and were sacrificed 17 days following pp lesion. Four groups of rats were tested: 1)MS-VDB with sham bilat-pp; 2)sham MS-VDB with bilat-pp; 3)MS-VDB with bilat-pp; 4)sham MS-VDB with sham bilat-pp. Using postembedding immunogold electron microscopy and SynBin, a program designed for quantification and compartmentalization of immunogold particles at the synaptic level, we investigated these effects in the outer molecular layer of the DG in a pilot study with 2 animals/group. Initial results suggest that the synaptic pools of NR1 within post-synaptic compartments were not affected with single MS/VDB, but that a long term synaptic down regulation of NR1 follows bilat pp lesion that is not affected by the additional removal of cholinergic input. While these combined lesions do not alter the pattern of synaptic NR1 receptor distribution following pp lesions, these data has important implications for lesion-induced hippocampal plasticity as well as structural and functional recovery.

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Yoder RM, Pang KCH (2003) Medial septal and entorhinal cortical involvement in hippocampal theta rhythm. Neuroscience 2003 Abstracts 719.19. Society for Neuroscience, New Orleans, LA.

Summary: Hippocampal theta rhythm (HPCΘ) may be involved in various phenomena, including attention and the acquisition of sensory information. Two projections to HPC, the medial septum-diagonal band of Broca (MSDB) and entorhinal cortex (EC), are involved in the activation or synchronization of HPCΘ. MSDB contains excitatory (cholinergic) and inhibitory (GABAergic) hippocampal projections via the fimbria/fornix. EC contains excitatory (glutamatergic) hippocampal projections via the perforant path (PP). MSDB GABAergic, MSDB cholinergic, or bilateral PP lesions eliminate HPCΘ during urethane anesthesia (HPCΘ-II). In unanesthetized recordings, each of these lesions reduced but did not eliminate HPCΘ during locomotion (HPCΘ-I); MSDB cholinergic and EC lesions caused similar reductions in HPCΘ, and MSDB GABAergic lesions produced a greater amplitude reduction. In an attempt to determine whether interactions exist between MSDB projections and EC, we examined the effects of MSDB GABAergic or cholinergic lesions combined with PP lesions on HPCΘ-I. MSDB GABAergic and cholinergic lesions were produced by intraseptal injection of kainic acid and 192 IgG-saporin, respectively. Bilateral PP lesions were produced by passing cathodal current through an electrode located in the medial PP. HPCΘ amplitude was calculated as the square root of power at peak frequency (Fourier analysis) within the HPCΘ range. The combination of MSDB GABAergic and PP lesions eliminated HPCΘ-I. The combination of MSDB cholinergic and PP lesions did not reduce HPCΘ-I amplitude further than MSDB cholinergic or PP lesions alone. These results suggest the inhibitory (MSDB GABAergic) and excitatory (MSDB cholinergic or EC glutamatergic) projections interact to support HPCΘ-I. Furthermore, MSDB cholinergic and EC glutamatergic projections may be redundant for HPCΘ-I.

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Fargo KN, Sengelaub DR (2003) Testosterone treatment protects motoneurons from dendritic atrophy following contralateral motoneuron depletion. Neuroscience 2003 Abstracts 602.2. Society for Neuroscience, New Orleans, LA.

Summary: In male rats, motoneurons of the spinal nucleus of the bulbocavernosus (SNB) project to the bulbocavernosus and levator ani muscles. SNB motoneurons and their target muscles are dependent on testosterone (T). We have previously demonstrated that unilateral depletion of SNB motoneurons induces dendritic atrophy in contralateral SNB motoneurons, and this atrophy is prevented by androgen manipulation. In the previous study, males were castrated for 6 weeks, then given replacement T coincident with motoneuron depletion. Because castration results in SNB dendritic retraction, and T replacement causes SNB dendrites to regrow to normal length, it is possible that the regressive changes or the active regrowth are involved in the protective effect of T manipulation. Alternatively, it may be that the effect can be accounted for simply by the high-normal levels of T produced by hormone implants. In the present experiment we show that SNB motoneuron dendrites are protected from contralateral motoneuron depletion by exogenous T alone (i.e., with no delay between castration and T replacement). We unilaterally depleted SNB motoneurons in male rats by intramuscular injection of cholera toxin conjugated saporin. Simultaneously, some saporin-injected rats were castrated and immediately given implants containing T. Four weeks later, contralateral SNB motoneurons were labeled with cholera toxin conjugated HRP, and dendritic arbors were reconstructed in 3 dimensions. A group of intact control males was also used. Contralateral SNB motoneuron depletion induced dendritic retraction to about 40% of normal length, but this atrophy was completely prevented by T treatment. Thus, the protective effect of T on SNB motoneurons is not due to prior dendritic retraction or T-induced regrowth per se. Instead, the presence of high-normal levels of T prevents dendritic retraction induced by contralateral motoneuron depletion.

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Chance WT, Dayal R, Friend LA, Sheriff S (2003) Normalization of burn-induced hypermetabolism following 3rd ventricle injection of saporin-CRF conjugate. Neuroscience 2003 Abstracts 614.5. Society for Neuroscience, New Orleans, LA.

Summary: The development of hypermetabolism following major burn trauma presents significant problems for patient management and recovery. To better understand CNS mediation of burn-induced hypermetabolism, we disrupted CRF neurotransmission by injecting a conjugate of CRF to the ribosome toxin, saporin, into the 3rd ventricle of burned and control rats. Following anesthetization (ketamine/xylazine; 80/15 mg/kg), cannulae (24 ga) were implanted into the 3rd ventricle of 58 adult, male, SD rats. Two weeks later, these rats were anesthetized again and subjected either to a 25 sec, open-flame, full-thickness burn or sham-burn procedures. One week later, the rats were divided into groups to receive ivt injections of artificial CSF (5 ul), saporin (2.5 ug) or saporin-CRF (2.5 ug). Resting energy expenditure (REE) was determined by indirect calorimetry for 60 min on all rats 14 days post-burn. Burned rats treated with CSF exhibited significantly (p<0.01) increased REE (164 ± 4 vs 132 ± 7 kcal/kg/day). Although the saporin treatment had no effect in burned (175 ± 6 kcal/kg/day) or sham-burned (133 ± 10 kcal/kg/day) rats, REE was reduced (p<0.01)in the burned rats treated with the saporin-CRF conjugate (145 ± 6 kcal/kg/day). CRF-induced (1 ug) increase in REE was also prevented in saporin-CRF-treated sham-burned rats. Determination of hypothalamic CRF receptor mRNA by RT-PCR suggested that CRF-R2 expression was reduced in saporin-CRF-treated rats, while CRF-R1 expression was not affected. These results suggest that hypothalamic CRF activity is involved in the maintenance of burn-induced hypermetabolism, and the CRF-2 receptor is important for the expression of this increase in REE. Therefore, control of hypermetabolism may be possible using selective CRF-R2 antagonists.

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