targeted-toxins

De Rosa E, Hasselmo ME, Baxter MG (2000) Contribution of the cholinergic basal forebrain to proactive interference between stored odor memories during associative learning in rats: 192 IgG-saporin immunotoxic lesions. Neuroscience 2000 Abstracts 563.8. Society for Neuroscience, New Orleans, LA.

Summary: Previous electrophysiological studies and a computational model suggest that cholinergic neuromodulation may reduce olfactory associative interference during learning (Hasselmo & Bower, 1993; Hasselmo et al., 1992). Using a simultaneous discrimination task where rats were required to learn a baseline odor pair (A+B-) and then two novel odor pairs: A-C+ (with an overlapping component A) and D+E- (with no overlapping component), De Rosa & Hasselmo (2000) demonstrated that a 0.25 mg/kg systemic dose of scopolamine (SCOP) selectively increased proactive interference. Under the influence of SCOP, the rats were impaired on acquiring the odor pair AC and not the odor pair DE relative to their normal saline performance. To localize this effect male Sprague-Dawley rats, with bilateral selective cholinergic lesions of the horizontal limb of the diagonal band of Broca (HDB group) or of all of the cholinergic nuclei of the basal forebrain (BF group), were tested on our task. Neither lesion impaired normal acquisition of either odor pair relative to the sham-operated control rats. However, the BF group, but not the HDB and control groups, were sensitive to a lower dose of SCOP (0.125 mg/kg) than in the previous study: this dose selectively impaired the BF group on acquiring the odor pair AC and not the odor pair DE relative to their normal saline performance, suggesting that weaker cholinergic modulation after removing the majority of cholinergic neurons in the basal forebrain makes the system more sensitive to proactive interference during blockade of remaining cholinergic effects.

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

Rogers SD, Salak-Johnson JL, Schwei MJ, Pomonis JD, Mantyh PW (2000) Reduced anxiety related behavior following ablation of amygdala neurons expressing substance P receptor. Neuroscience 2000 Abstracts 571.2. Society for Neuroscience, New Orleans, LA.

Summary: The neurokinin substance P (SP) is localized in brain regions that coordinate stress response and may play a role in modulating anxiety. Effects of ablation of substance P receptor (SPR)-expressing neurons by administration of a substance P-toxin conjugate, substance P-saporin (SP-SAP), in the amygdala (a brain region known to modulate stress and anxiety responses) were examined immunohistochemically and behaviorally thirty days following SP-SAP treatments. Rats were bilaterally injected in basolateral amygdala nuclei with 5μl of sterile saline, 1 μM saporin (SAP), or 1 μM SP-SAP. SPR-immunofluoresence levels and number of SPR-IR positive neurons in amygdalar subnuclei decreased following SP-SAP treatment. SP-SAP did not induce significant gliosis or non-specific neuronal death. Interestingly, after SP-SAP treatment, the number of NPY-IR neurons were also decreased, and combined SPR and NPY immunofluorescence demonstrated a large number of NPY-IR neurons colocalize with SPR-IR neurons in the amygdala. Thirty days following SP-SAP treatment, rats were tested in elevated plus maze (EPM) and open field (OF). Anxiety level and exploratory behavior displayed by SP-SAP treated rats were altered; they had significantly more entries into and spent more time in EPM open arms than did saline- or SAP-injected rats. In the OF, SP-SAP treated rats spent less time frozen than saline or SAP treated rats. These results suggest that SPR expressing neurons in the amygdala plays a pivotal role in generation of anxiety behaviors and that SP may play a modulatory role in stress-induced anxiety behavior.

Related Products: SP-SAP (Cat. #IT-07)

McGaughy JA, Dalley JW, Robbins TW, Everitt BJ (2000) The behavioral and neurochemical effects of acute and incremental cholinergic lesions on visual attention during a 5-choice serial reaction time task. Neuroscience 2000 Abstracts 563.1. Society for Neuroscience, New Orleans, LA.

Summary: Previously studies have shown that infusions of 192 IgG-saporin (SAP)into the nucleus basal magnocellularis (nbm) produce severe,persistent impairments in visual attention that correlate with the extent of cortical cholinergic deafferentation. After one bilateral infusion of a low dose (0.15 μg/μl; LX1) of SAP into the nbm, rats performing in a 5 CSRTT show specific behavioral impairments when tested with an increased event rate (ITI=2sec). In vivo microdialysis performed while rats were tested under standard conditions (ITI=5sec) in the 5CSRTT showed that LX1 rats had an increased basal acetylcholine (ACH) efflux in the prefrontal cortex (PFC) relative to sham-lesioned animals on the first day of testing, but no difference in task related efflux on either day of testing. The current study investigated the effects of larger cholinergic lesions on attentional perfomance to determine if they would produce large decreases in cortical ACH efflux and consequently impair attention. Rats were trained in a 5 CSRTT prior to receiving intra-nbm infusions of SAP or vehicle (VEH). Lesioned animals were subjected to repeated infusions of a low dose of the toxin (0.15 μg/μl), a single high dose (0.45 μg/μl) or vehicle. The (0.45 μg/μl) dose of SAP produced profound impairments under baseline conditions that correlated with low levels of ACH efflux in the PFC. These impairments were not attenuated by systemic nicotine or physostigmine. The cumulative lesions were hypothesized to increase sensitivity to attentional demands and pharmacological intervention.

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

Janni G, Moallem T, Lappi DA, Ohara PT, Jasmin L (2000) Schwann cells are removed from the rat spinal cord after effecting recovery from paraplegia. Neuroscience 2000 Abstracts 516.8. Society for Neuroscience, New Orleans, LA.

Summary: Remyelination of the CNS is necessary to restore neural function in a number of demyelinating conditions such as multiple sclerosis. Schwann cells, the myelinating cells of the periphery, are good candidates for this purpose, having more robust regenerative properties than their central homologues, the oligodendrocytes. While the ability of Schwann cells to remyelinate the CNS and effect functional recovery has been demonstrated, their long term survival in the CNS after myelinating central axons is largely unknown. We use saporin conjugated to the cholera toxin B-subunit to demyelinate the rat lumbar spinal cord, remove macroglia, and produce paraplegia. This treatment is followed by a spontaneous proliferation of large numbers of endogenous Schwann cells which remyelinate spinal cord axons with concomitant functional recovery from paraplegia within 75 days. During the following weeks, however, quantification on thin sections shows that Schwann cells are progressively replaced by oligodendrocytes, without any lapse in behavioral recovery. This removal of Schwann cells is confirmed by ultrastructural examination and by immunocytochemistry for Schwann cells and oligodendrocytes. Our results indicate that Schwann cell remyelination of the spinal cord might not be permanent. They can be induced to demyelinate and desheath through endogenous mechanisms that remain to be characterized.

Related Products: CTB-SAP (Cat. #IT-14)

Winters BD, Dunnett SB (2000) Combined cholinergic denervation of the hippocampus and posterior cingulate cortex fails to impair working memory performance but may produce deficits in behavioural flexibility in the rat. Neuroscience 2000 Abstracts 563.2. Society for Neuroscience, New Orleans, LA.

Summary: Selective cholinergic denervation of either the hippocampal formation (HPC) or posterior cingulate cortex (pCCX) with the immunotoxin 192 IgG-saporin produces negligible effects on rats’ performance of the delayed nonmatching-to-position (DNMTP) task. Yet, fimbria-fornix transection, which disrupts the cholinergic input to both of these limbic regions, causes a delay-dependent deficit in this working memory task. In the current study, rats were trained in a standard DNMTP procedure and then divided into three groups: Group SAP (n=6) received injections of 192 IgG-saporin into both HPC and pCCX; Group NMDA (n=4) received similar injections of the excitotoxin N-methyl-D-aspartic acid; and Control rats (n=10) received vehicle injections. Following surgery, NMDA rats performed significantly worse than SAP and Control rats at all delays in the DNMTP task (p<0.05); SAP rats did not differ from Controls. All groups then acquired a matching-to-position task (i.e., reversal) over four sessions; however, SAP rats performed significantly more perseverative errors during the first reversal session, resulting in a significant Group x Session interaction (p<0.01). It is suggested that, while the cholinergic projections to the HPC and pCCX are not crucial for working memory performance in the DNMTP task, the cholinergic innervation of these limbic regions may influence behavioural flexibility by modulating a circuit mediating habit-like performance.

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

Blanco-Centurion CA, Salin-Pascual R, Gerashchenko D, Greco MA, Lappi DA, Kilduff TS, Shiromani PJ (2000) DBH-saporin lesions the locus coeruleus, but does not produce cataplexy or abnormal REM sleep triggering. Neuroscience 2000 Abstracts 566.17. Society for Neuroscience, New Orleans, LA.

Summary: Recently, canine narcolepsy was associated with a mutation in the hypocretin-2 receptor (Lin et al., 1999), which binds the neuropeptide hypocretin, also known as orexin. The locus coeruleus receives a very heavy projection of HCRT/OX fibers, and the LC also contains HCRT/OX receptor mRNA. Silence of LC neurons is hypothesized to be key in triggering cataplexy and REM sleep. To test this hypothesis, DBH-saporin was used to selectively lesion the LC. Male Sprague-Dawley rats (400-450 g) instrumented for recording sleep were given DBH-saporin (n=4) (500ng/0.5ul) via a micropipette to the LC. Control rats were administered 192-saporin (n=3), or saline (n=4). Two days later, sleep recordings were obtained for 7 consecutive days. The rat’s behavior was videotaped at night. To identify whether cataplexy was induced, the alpha antagonist, Prazosin was administered (1500h, 500mg/kg, IP) and then sleep and video recordings were made for three hours. Brains were removed for histology. DBH-saporin completely lesioned the LC neurons. However, there were no changes in wakefulness, nonREM or REM sleep. Video recordings also did not reveal any cataplexy episodes. The application of Prazosin did not induce cataplexy or diminish muscle tone in DBH-saporin LC treated rats. Historically, LC lesions have never been found to induce cataplexy. Use of DBH-saporin provides a more specific lesion restricted to the LC neurons where the HCRT/OX receptor mRNA is localized. Our findings indicate that LC neurons are not essential for maintaining muscle tone or wakefulness.

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

Schreihofer AM, Stornetta RL, Guyenet PG (2000) Regulation of sympathetic vasomotor tone and arterial pressure by the rostral ventrolateral medulla after elimination of C1 neurons in rat. Neuroscience 2000 Abstracts 310.7. Society for Neuroscience, New Orleans, LA.

Summary: The rostral ventrolateral medulla (RVLM) tonically stimulates sympathetic preganglionic neurons to maintain arterial pressure (AP). Although the C1 neurons in the RVLM may have a sympathoexcitatory function, it is not known whether they are the essential presympathetic RVLM neurons. In the present study, we selectively destroyed spinally projecting C1 cells (∼84%) with bilateral microinjections (spinal segments T2-T3) of an anti-dopamine-betahydroxylase antibody conjugated to saporin (anti-DβH-SAP). 3-5 weeks later these rats had a normal AP and splanchnic nerve activity (SNA) under chloralose anesthesia. Extracellular recording and juxtacellular labeling of bulbospinal barosensitive neurons in RVLM revealed that after anti-DβH-SAP only the lightly myelinated RVLM neurons with no or very low levels of tyrosine hydroxylase immunoreactivity were preserved. In these rats, inhibition of RVLM (muscimol 100 pmol/100 nl/side) eliminated SNA and decreased AP as seen in control rats. However, treatment with anti-DβHSAP reduced the sympathoexcitatory and pressor responses to electrical stimulation in RVLM. Although treatment with anti-DβH-SAP also eliminated A5 noradrenergic cells, rats with selective lesions of A5 cells (local microinjection of 6-hydroxydopamine) displayed no deficits to stimulation of the RVLM. These data suggest basal sympathetic vasomotor tone relies primarily on non-catecholaminergic presympathetic cells in the RVLM. In contrast, bulbospinal adrenergic neurons are important for the increased SNA and AP produced by stimulation of the RVLM.

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

Ciriello J, Rosas-Arellano MP, Solano-Flores LP (2000) Intrathecal injections of saporin conjugated substance-P alter the development of renal hypertension. Neuroscience 2000 Abstracts 310.2. Society for Neuroscience, New Orleans, LA.

Summary: We have recently shown that there is an up-regulation of mRNA encoding Substance-P (SP) in dorsal root ganglia in the two kidney, one clip (2K1C) model of renal hypertension. In this study, the effect of intrathecal injections of the toxin saporin conjugated to SP (SAP; 5 x 10-6 M) on the development of 2K1C hypertension was investigated in male Wistar rats. Rats were randomly assigned to 2 groups and instrumented with an intrathecal cannula that ended at the T10-T12 spinal level. Arterial pressure (AP) and heart rate (HR) were recorded using the indirect tail-cuff method. After a 1 week control period, one group of animals received a single 10 µl injection of SAP and the other an equal volume of the saline vehicle (control). Two weeks later a clip was placed on the left renal artery in all animals. Injections of either SAP or the vehicle did not alter AP and HR compared to pre-injection levels. In addition, AP and HR were not altered in the SAP group compared to controls for the 2 week period prior to the renal clip. However, during the 4 weeks after the renal clip, AP and HR in the SAP group were significantly lower compared to the control animals. These data suggest that dorsal horn neurons containing SP receptors and that receive afferent renal nerve inputs are involved in renal hypertension.

Related Products: SP-SAP (Cat. #IT-07)

Vulchanova L, Olson TH, Elde R, Honda CN (2000) Role of IB4-binding sensory neurons in the effects of intradermal capsaicin injection. Neuroscience 2000 Abstracts 212.7. Society for Neuroscience, New Orleans, LA.

Summary: We have shown previously that a unilateral injection of a conjugate of the lectin IB4 and the toxin saporin (IB4-SAP) into sciatic nerve of rats results in loss of IB4-binding neurons and transient increase in thermal and mechanical nociceptive thresholds. The thresholds were maximally increased 10 days post-treatment and returned to baseline levels by day 21. In the present study, we examined the responses of IB4-SAP treated rats after intradermal injection of capsaicin, which results in acute nocifensive behavior followed by thermal and mechanical hyperalgesia. The nocifensive behavior of IB4-SAP treated rats 10, 21 and 42 days post-treatment was 6%, 36% and 47%, respectively, of the behavior of control treated rats. IB4-SAP treated rats injected with capsaicin did not develop thermal or mechanical hyperalgesia at any of the time points examined. These results suggest that the increase in thermal nociceptive thresholds after IB4-SAP treatment is due to loss of VR1-expressing IB4-binding neurons since the nocifensive behavior is most likely mediated by the capsaicin receptor VR1, which also transduces noxious thermal stimuli. In addition, VR1 in surviving neurons may contribute to the recovery of thermal nociceptive thresholds. Finally, our results suggest that IB4-binding neurons are required for development of capsaicin-mediated hyperalgesia, and that the recovery of the responsiveness of IB4-SAP treated rats to noxious stimuli under normal conditions is not accompanied by recovery of the mechanisms underlying hyperalgesia.

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

Martin JL, Sloviter RS (2000) Focal hippocampal hyperexcitability after focal interneuron ablation in the rat by substance P-saporin. Neuroscience 2000 Abstracts 389.13. Society for Neuroscience, New Orleans, LA.

Summary: Hyperexcitability after prolonged seizures or head trauma may result from interneuron malfunction or loss; but a causal relationship is in doubt because global insults produce widespread brain damage and other effects. We have therefore sought to destroy interneurons selectively using stable Substance P-saporin (SSPsap; ATS); a neurotoxin internalized by SP receptor (SPR)-expressing neurons. Improved immunofluorescent methods revealed that most GABA-; parvalbumin (PV)-; and somatostatin (SS)-positive (+) cells of all hippocampal regions (dentate gyrus and areas CA1-CA3) are SPR+; but that granule cells; mossy cells; and CA1-3 pyramidal cells are not. Intrahippocampal injections of SSPsap or vehicle were made under urethane anesthesia in 3 sites (20nl/site) of the dorsal hippocampus of 6 male Sprague-Dawley rats/group. After 5-90 days; rats were blindly evaluated in two sites for CA1 pyramidal cell and dentate granule cell responses to perforant path stimulation (PPS). SSPsap-treated rats exhibited relatively normal responses in some sites; but pathophysiology at other sites that was virtually identical to that seen after prolonged PPS or kainate (multiple population spikes and paired-pulse disinhibition in response to 0.1-2.0Hz perforant path (PP) stimuli). Abnormal responses were observed at the earliest time tested (5 days); and at 90 days. Anatomical analysis revealed selective loss of SPR+; PV+; SS+; and GABA+ neurons; and survival of principal cells and extrinsic afferents. Importantly; “epileptic” pathophysiology was observed exclusively in areas of interneuron loss. These data indicate that the pathophysiology produced by status epilepticus or head trauma can be replicated focally by selective interneuron loss alone; and provide the first direct evidence that highly focal interneuron loss per se is capable of replicating “epileptic” disinhibition and hyperexcitability. In addition; the pathophysiology is restricted to the region of the affected interneuron somata; suggesting a highly localized influence of inhibitory interneurons. Supported by: NIH grant NS18201.

Related Products: SSP-SAP (Cat. #IT-11)

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