targeted-toxins

Parker RA, Wang F, Hayashida K, Martin TJ, Eisenach JC, Peters CM (2013) Descending noradrenergic control of conditioned pain modulation and postoperative pain trajectory in rats. Neuroscience 2013 Abstracts 461.10. Society for Neuroscience, San Diego, CA.

Summary: Aim of Investigation: Chronic pain after surgery (CPAS) is now recognized as a significant clinical problem that occurs in 10-50% of patients undergoing surgery. Recent clinical studies demonstrate the integrity of endogenous pain inhibitory controls are important for preventing CPAS, however this relationship and underlying mechanisms haven’t been examined in preclinical models. We hypothesized that a causal relationship exists between impaired endogenous analgesia and chronic pain after surgery that is in part dependent on descending noradrenergic pathways. Methods: We examined the integrity of endogenous analgesia in rats preoperatively used a previously described method for assessing conditioned pain modulation (CPM) in rats (Ferrari et al., 2010) involving injection of capsaicin (150μg/50μl) in the forepaw as a conditioning stimulus and detection of hindpaw mechanical thresholds (Randall-Selitto device) as the test stimulus. The partial L5 spinal nerve ligation (pSNL) model and mixed effects growth curve modeling of mechanical withdrawal thresholds assessed for 10 weeks following surgery were used to study resolution of postoperative mechanical hypersensitivity. The role of descending spinal noradrenergic pathways in CPM and postoperative trajectory was assessed by ablating spinal noradrenergic fibers with DβH-saporin or blocking their activity pharmacologically with adrenergic receptor antagonists. Results: Forepaw capsaicin resulted in release of NE in the lumbar spinal cord and this CPM paradigm was partially prevented by spinal idazoxan (30μg, i.t.). CPM assessed preoperatively resulted in pronounced but variable increases in hindpaw mechanical withdrawal thresholds at 30 minutes (range: 60-140g). Within individual rats, we observed a significant correlation between the degree of preoperative CPM and the slope of trajectory (P=0.006, r=0.610) as rats with lower endogenous analgesia had slower resolution of mechanical hypersensitivity. In support of a causal role between endogenous spinal noradrenergic activity and CPAS, depletion of spinal noradrenergic fibers prior to pSNL resulted in a significant reduction in the slope of trajectory within the ipsilateral hindpaw. Conclusions: Collectively, these studies suggest that the ability to engage descending endogenous noradrenergic pathways may be critical in determining whether CPAS develops. Furthermore, the use of growth curve modeling to study CPAS will allow us to examine the ability of socio-environmental conditions or pharmacological interventions to impair or improve aspects of postoperative pain trajectory as part of future studies.

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

Pinto LG, Souza GR, Lopes AHP, Talbot J, Cunha FQ, Cunha TM, Ferreira SH (2013) Role of nonpeptidergic subset of primary afferent neurons in inflammatory hypernociception in mice. Neuroscience 2013 Abstracts 256.15. Society for Neuroscience, San Diego, CA.

Summary: Sensory information is transmitted from the periphery to the spinal cord by distinct subsets of primary afferent neurons, including small diameter unmyelinated C-fibers, which plays an important role in detecting noxious stimuli. C-fiber nociceptors have been divided into two classes, the peptidergic and nonpeptidergic. While many of the differences between peptidergic and nonpeptidergic neurons are now appreciated, a possible functional difference between these two classes of C fibers in the genesis of acute nociception as well as inflammatory pain is still unclear. Thus, this study aims to clarify the role of nonpeptidergic C fibers in acute nociception induced by mechanical, thermal and chemical stimuli as well as in inflammatory hypernociception. In order to elucidate differences between these two classes of C fibers, a neurotoxin was used to selectively eliminate the nonpeptidergic C fibers: a saporin conjugated to isolectin B4 (IB4). Nociceptive threshold was evaluated through thermal (Hargreaves) and mechanical (filaments and electronic von Frey) tests in C57BL/6 mice. Nociception models were induced by intraplantar (i.pl.) injection of capsaicin and formalin (acute nociception) or by i.pl. administration of prostaglandin E2 (PGE2), epinephrine, endothelin, NGF, GDNF and carrageenan (inflammatory hypernociception). P2X3 and TRPV1 expression were analyzed by Western blot of dorsal root ganglion (DRG). The expression of IB4-labeled in spinal cord was determined by immunofluorescense using confocal microcopy. Firstly, it was observed that the intrathecal administration of IB4-saporin did not change baseline thermal and mechanical nociceptive threshold of the mice paw when compared to saline and saporin-control groups. The intrathecal administration of IB4-saporin reduced mechanical inflammatory hypernociception induced by carrageenan, epinephrine, endothelin, PGE2 or GDNF, but not NGF, in mice. Similarly, the treatment with IB4-saporin inhibited the nociception caused by intraplantar injection of the capsaicin. By contrast, the acute nociception induced by formalin did not change by administration of IB4-saporin. In addition, the expression of TRPV1 and P2X3 in DRG were reduced after treatment with IB4-saporin. Consistent with these findings, we found that IB4-saporin injection decreased the expression of IB4-labeled in spinal cord. These results suggest that absence the nonpeptidergic C fibers does not affect basal nociceptive threshold. However, these fibers are essential for the development of nociception in the paw of mice induced by inflammatory stimuli like PGE2, epinephrine, endothelin, carrageenan and capsaicin.

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

Krajewski-Hall SJ, Mittelman-Smith, Mcmullen NT, Rance NE (2013) Ablation of arcuate KNDy neurons amplifies the LH surge in steroid-primed, ovariectomized rats. Neuroscience 2013 Abstracts 274.01. Society for Neuroscience, San Diego, CA.

Summary: KNDy (kisspeptin, neurokinin B and dynorphin) neurons in the arcuate nucleus play an important role in the reproductive axis. We have developed a method to selectively ablate KNDy neurons in the rat using NK3-SAP, a neurokinin 3 receptor agonist conjugated to saporin (Mittelman-Smith, Endocrinology 2012). Ablation of KNDy neurons results in cessation of estrous cycles, ovarian atrophy, a decrease in tonic LH secretion and loss of the rise of serum LH after ovariectomy. Given these profound effects, we tested if we could induce an LH surge in KNDy-ablated rats using a well-established steroid replacement regimen. Rats were maintained on a 14:10 light cycle (lights on at 0500). Using stereotactic surgery, NK3-SAP or Blank-SAP was injected in the arcuate nucleus and rats were allowed to recover for one month before ovariectomy. Seven days after ovariectomy they were implanted with silastic capsules containing 17β-estradiol. Two days later, they were implanted with progesterone capsules (~0830h). Rats were sacrificed in the afternoon at a time previously shown to exhibit peak LH surge levels (~1600h) and the brains were processed for immunohistochemistry. Ablation of KNDy neurons was verified by near complete loss of NKB-immunoreactive neurons in the arcuate nucleus in NK3-SAP rats. At 0830h, tonic levels of serum LH were significantly lower in KNDy ablated rats, consistent with our previous studies. Unexpectedly, the surge in serum LH at 1600h was more than 3-fold higher in NK3-SAP-treated rats compared to Blank-SAP controls (53.5 + 16.5 ng/ml vs 16.5 + 2.1 ng/ml, respectively). To determine if this change was associated with increased activation of GnRH neurons, dual-labeled GnRH-fos immunocytochemistry was performed in rostral hypothalamic sections. There was no significant difference in the total number of GnRH cells counted in 4 matched sections (NK3-SAP, 85.8 + 9.8 vs Blank-SAP, 84.9 + 4.6) or in the percentage of GnRH cells that were activated, as measured by GnRH-fos coexpression (NK3-SAP, 22.4 + 1.8% vs Blank-SAP, 16.6 + 4.9%). In addition, there was no difference between NK3-SAP and Blank-SAP controls in the number of fos-ir cells counted in the AVPV. These data indicate that arcuate KNDy neurons are not required for the induction of an LH surge. The marked increase in the LH surge in KNDy-ablated rats, however, suggests that KNDy neurons are important for regulating the magnitude of the surge.

Related Products: NKB-SAP (Cat. #IT-63)

Burk JA, Yonezaki K (2013) Loss of medial prefrontal cortical cholinergic projections increases preference for an immediately available reward in a delay discounting task. Neuroscience 2013 Abstracts 288.02. Society for Neuroscience, San Diego, CA.

Summary: Attentional control is thought to regulate numerous processes, including impulsive choices. The cholinergic projections to the medial prefrontal cortex are thought to be part of a distributed neural circuit that maintains attentional control. In the present experiment, male FBNF1 hybrid rats were trained in a delay discounting task that involved a choice between a small, immediately available reward and a larger reward. The larger reward was also immediately available at the beginning of the test session and then the delay to receive this reward was progressively increased within a session (0-40 s). After reaching stable performance levels, rats then either received infusions of the cholinergic immunotoxin, 192IgG-saporin, or vehicle into the medial prefrontal cortex. After recovering from surgery, rats were tested in the same delay discounting task that had been trained prior to surgery. Relative to sham-lesioned animals, rats with a loss of the cholinergic projections to the medial prefrontal cortex exhibited an increased preference for selecting the smaller, immediate reward. Subsequent administration of nicotine (0.0, 0.1, 0.2, 0.4 mg/kg, ip) did not substantially alter the effects of the lesion on delay discounting performance. The present results suggest that medial prefrontal cortical cholinergic projections contribute to choice behavior based upon delay to reward access and reward magnitude. Moreover, these results are consistent with the idea that disruption of attentional control can increase impulsive choices.

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

Lee J, Jeong D, Oh J, Lee J, Chang W, Cho Z, Chang J (2013) Selective basal forebrain cholinergic deficits reduce glucose metabolism, cholinergic and GABAergic system in the cingulate cortex. Neuroscience 2013 Abstracts 45.12. Society for Neuroscience, San Diego, CA.

Summary: Reduction of brain glucose metabolism and degeneration of cholinergic basal forebrain neurons are common features in Alzheimer’s disease and have been correlated with memory function. Although regions representing glucose hypometabolism in Alzheimer’s disease patients are target sites of cholinergic basal forebrain neurons, an interaction between cholinergic denervation and glucose hypometabolism is still unclear. To evaluate the changes in glucose metabolism in regions relevant to basal forebrain cholinergic deficits, we damaged basal forebrain cholinergic neurons of rats using 192 IgG-saporin. After 3 weeks, lesioned animals were tested by water maze test or analyzed using 18F-2-fluoro-2-deoxyglucose positron emission tomography. During the probe test in the water maze, performance of the lesion group decreased, considering time spent in both the target quadrant and platform zone. Glucose metabolism in the cingulate cortex of the lesion group decreased compared with the normal group. Additionally, acetylcholinesterase activity and glutamate decarboxylase 65/67 expression both declined in the cingulate cortex. Our results reveal that spatial memory impairment of animals in which basal forebrain cholinergic neurons are selectively damaged is associated with a decline in functions of GABAergic, cholinergic, and glutamatergic systems associated with glucose hypometabolism in the cingulate cortex.

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

Wiese AJ, Rathbun M, Butt MT, Malkmus SA, Richter PJ, Osborn KG, Xu Q, Veesart SL, Steinauer JJ, Higgins D, Lappi DA, Russell B, Yaksh TL (2013) Intrathecal substance p-saporin in the dog: distribution, safety, and spinal neurokinin-1 receptor ablation. Anesthesiology 119(5):1163-1177. doi: 10.1097/ALN.0b013e3182a95164

Summary: Here the authors investigate the safety parameters of SP-SAP on purpose-bred beagles (currently in human clinical trials). The dogs received 1.5, 15, or 150 μg intrathecal injections of the conjugate. SP-SAP pharmacology and physiological effects were assessed by behavioral and functional observations, immunohistochemistry, ELISA, blood and urine analysis, histopathology, and in situ hybridization. The general conclusions include that neurokinin-1 receptor (NK1r) positive neuron loss is detectable as soon as 7 days after administration of SP-SAP, the neuron loss is permanent, toxicity is specific to NK1r-positive neurons, and, other than the 150 μg dose, NK1r neuron loss was restricted to the superficial dorsal horn.

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

Read the featured article in Targeting Trends.

Brown DC, Agnello K (2013) Intrathecal substance p-saporin in the dog: efficacy in bone cancer pain. Anesthesiology 119(5):1178-1185. doi: 10.1097/ALN.0b013e3182a95188

Summary: This work demonstrates the use of naturally occurring bone cancer in dogs as a model for pain therapy. Companion dogs with bone cancer received 20-60 μg intrathecal injections of SP-SAP (currently in human clinical trials) depending on the size of the dog. Significantly more dogs in the control group required unblinding and adjustment of pain care than in the SP-SAP group, indicating the efficacy of SP-SAP in pain control. This study also demonstrates the validity of the dog model for testing analgesic protocols.

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

Read the featured article in Targeting Trends.

Hayashida K (2013) Substance P-saporin for bone cancer pain in dogs: Can man’s best friend solve the lost in translation problem in analgesic development?. Anesthesiology 119(5):999-1000. doi: 10.1097/ALN.0b013e3182a951a2

Summary: This editorial describes the SP-SAP papers in this latest issue of Anesthesiology. The results of the paper are discussed, and the potential in using companion dogs for pain models is emphasized. While most pain models have been rodent-based, companion dogs provide models for chronic pain due to natural causes such as cancer and arthritis, along with frequent opportunity for behavioral assessments by the owner. Such assessments can be done without stress to the animal.

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

See Also:

• Wiese AJ et al. Intrathecal substance p-saporin in the dog: distribution, safety, and spinal neurokinin-1 receptor ablation. Anesthesiology 119(5):1163-1177, 2013.
• Brown DC et al. Intrathecal substance p-saporin in the dog: efficacy in bone cancer pain. Anesthesiology 119(5):1178-1185, 2013.

Laplante F, Dufresne MM, Ouboudinar J, Ochoa-Sanchez R, Sullivan RM (2013) Reduction in cholinergic interneuron density in the nucleus accumbens attenuates local extracellular dopamine release in response to stress or amphetamine. Synapse 67(1):21-29. doi: 10.1002/syn.21612

Summary: The authors examined whether excessive dopamine neurotransmission in the mesolimbic system is due to higher levels of presynaptic or postsynaptic dopamine. Rats received 250-ng bilateral injections of anti-ChAT-SAP (Cat. #IT-42) into the nucleus accumbens. Rabbit IgG-SAP (Cat. #IT-35) was used as a control. The data suggest that reduction of cholinergic interneurons in the nucleus accumbens suppresses presynaptic dopamine release.

Related Products: Anti-ChAT-SAP (Cat. #IT-42), Rabbit IgG-SAP (Cat. #IT-35)

Read the featured article in Targeting Trends.

Cutuli D, De Bartolo P, Caporali P, Tartaglione AM, Oddi D, D’Amato FR, Nobili A, D’Amelio M, Petrosini L (2013) Neuroprotective effects of donepezil against cholinergic depletion. Alzheimers Res Ther 5(5):50. doi: 10.1186/alzrt215

Summary: Acetylcholinesterase inhibitors appear to be one of the only pharmacological tools available to reduce cognitive deficits caused by the loss of cholinergic neurons in the basal forebrain. Here the authors pre-treated rats with the aceytlcholinesterase inhibitor donepezil before administering 0.5 μg of 192-IgG-SAP (Cat. #IT-01) into each side of the medial septum. Analysis of working memory, spatial discrimination, social novelty preference, and ultrasonic localizations, along with measuring hippocampal and neocortical caspase-3 activity indicates that donepezil pre-treatment ameliorates some effects of the cholinergic depletion.

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