2020 Scientific References

Targeted Toxins: Targeting agent attached to Saporin

IT-01 192-IgG-SAP

Intracerebroventricular Administration of 192IgG-Saporin Alters the State of Microglia in the Neocortex. Volobueva MN, Dobryakova YV, Manolova AO, Stepanichev MY, Kvichansky AA, Gulyaeva NV, Markevich VA, Bolshakov AP Neurochem J 14(1):37-42, 2020.  doi: 10.1134/S1819712420010213

Objective: The effect of intracerebroventricular immunotoxin administration on the state of microglia in tissues adjacent to the ventricle (striatum and parietal cortex) and remotely located but receiving innervation from the medial septal region and diagonal band of Broca (entorhinal cortex and olfactory bulbs). Summary: 1.5 months after the administration of immunotoxin, microglia are activated only in the neocortical areas, not in the striatum or olfactory bulbs. Dose: Injected bilaterally at a dose of 4 μg in each ventricle.

Long-term potentiation in the hippocampal CA3 to CA1 synapses may be induced in vivo by activation of septal cholinergic inputs. Dobryakova YV, Stepanichev MY, Markevich VA, Bolshakov AP Int J Neurosci 23:1-7, 2020.  doi: 10.1080/00207454.2020.1822834

Evaluation of the Adverse Effects of Chronic Exposure to Donepezil (An Acetylcholinesterase Inhibitor) in Adult Zebrafish by Behavioral and Biochemical Assessments. Audira G, Ngoc Anh NT, Ngoc Hieu BT, Malhotra N, Siregar P, Villalobos O, Villaflores OB, Ger TR, Huang JC, Chen KH, Hsiao CD Biomolecules 10(9):1340, 2020.  doi: 10.3390/biom10091340

Objective: The authors use zebrafish to conduct a deeper analysis of the potential adverse effects of DPZ on the short-term memory and behaviors of normal zebrafish by performing multiple behavioral and biochemical assays. Summary: Chronic waterborne exposure to DPZ can severely induce adverse effects on normal zebrafish in a dose-dependent manner. (Citation: Kimura, M.; Akasofu, S.; Ogura, H.; Sawada, K. Protective effect of donepezil against Aβ (1–40) neurotoxicity in rat septal neurons. Brain Res. 2005, 1047, 72–84.)

Acetylcholine and Spontaneous Recognition Memory in Rodents and Primates. Easton A, Barros M, Lever C Curr Top Behav Neurosci 45:29-45, 2020.  doi: 10.1007/7854_2020_132

Review of lesioning designed to specifically target cells that express acetylcholine as a transmitter.

Cholinergic Signaling Dynamics and Cognitive Control of Attention. Parikh V, Bangasser DA Shoaib M., Wallace T (Ed.): Behavioral Pharmacology of the Cholinergic System. Current Topics in Behavioral Neurosciences. 45:71-87, 2020.  Springer, Cham doi: 10.1007/7854_2020_133

Summary: A plethora of studies conducted in rodents demonstrated that selective lesions of BF cholinergic neurons and their cortical inputs produced by the immunotoxin 192-IgG-SAP impair performance in various tasks of attention.

Measuring attention in rats with a visual signal detection task: Signal intensity vs. signal duration. Holloway Z, Koburov R, Hawkey A, Levin ED Pharmacol Biochem Behav 199:173069, 2020.  doi: 10.1016/j.pbb.2020.173069

Summary: Drug-induced effects have been used to demonstrate the construct validity of operant attention tasks, as well as to assess the pharmacological systems that underlie cognitive processes, such as attention, short-term memory and reaction time, either by interrupting or enhancing performance. The authors reference: McGaughy J, Sarter M (1998) Sustained attention performance in rats with intracortical infusions of 192 IgG-saporin-induced cortical cholinergic deafferentation: effects of physostigmine and FG 7142. Behav Neurosci 112:1519-1525. doi: 10.1037/0735-7044.112.6.1519

Extracellular levels of the sleep homeostasis mediator, adenosine, are regulated by glutamatergic neurons during wakefulness and sleep. Sun MJ, Tang Y Purinergic Signal 16(4):475-476, 2020.  doi: 10.1007/s11302-020-09758-3

Summary: Blanco-Centurion et al. investigated the role of cholinergic neurons in the BF by administering 192–IgG–SAP to lesion them, but surprisingly the results indicated that adenosine from cholinergic neurons in BF are not essential to sleep induction. See: Blanco-Centurion C, Xu M, Murillo-Rodriguez E, Gerashchenko D, Shiromani AM, Salin-Pascual RJ, Hof PR, Shiromani PJ (2006) Adenosine and sleep homeostasis in the basal forebrain. J Neurosci 26(31):8092–8100.

Identification of Multiple Targets in the Fight against Alzheimer’s Disease. Giannoni P, Fossati S, Claeysen S, Marcello E, eds Front Aging Neurosci 12:169, 2020.  doi: 10.3389/fnagi.2020.00169

A collection of 20 articles that depict a broad representation of the most impactful advances in Alzheimer’s disease (AD) comprehension and therapeutic openings.

IT-02 OX7-SAP, discontinued

Reversal of Object Recognition Memory Deficit in Perirhinal Cortex-Lesioned Rats and Primates and in Rodent Models of Aging and Alzheimer’s Diseases. Masmudi-Martín M, Navarro-Lobato I, López-Aranda MF, Browning PGF, Simón A-M, López-Téllez JF, Jiménez-Recuerda I, Martîn-Montañez E, Pérez-Mediavilla A, Frechilla D, Baxter MG, Khan ZU Neuroscience 448:287-298, 2020.  doi: 10.1016/j.neuroscience.2020.08.039

IT-03 Anti-DBH-SAP

An age- and sex-dependent role of catecholaminergic neurons in the control of breathing and hypoxic chemoreflex during postnatal development. Patrone LGA, Capalbo AC, Marques DA, Bícego KC, Gargaglioni LH Brain Res 1726:146508, 2020.  doi: 10.1016/j.brainres.2019.146508

Objective: To discover the role of brainstem catecholaminergic (CA) neurons in the hypoxic ventilatory response (HVR). Summary: Brainstem CA neurons modulate the HRV during the postnatal phase, and possibly thermoregulation during hypoxia. Dose: Evaluation of brainstem CA neurons in the HVR during postnatal development  in male and female rats through specific cell depletion with Anti-DBH-SAP (420 ng/nL) injected in the fourth ventricle.

Integration of Peripheral and Central Systems in Control of Ingestive and Reproductive Behavior. Schneider J Oxford Research Encyclopedia of Neuroscience , 2020.  Oxford University Press doi: 10.1093/acrefore/9780190264086.013.23

Summary: Highly glucose-sensitive cells in the ventrolateral medulla send catecholaminergic projections to the PVH. These projections can be selectively destroyed by Anti-DBH-SAP “DSAP” lesions, wherein a toxin (saporin) is directed at nondopaminergic cells that express the enzyme necessary for synthesis of epinephrine and norepinephrine (the enzyme is dopamine beta hydroxylase). DSAP experiments show that catecholaminergic projections from glucose-sensitive cells in the ventrolateral medulla are necessary for all responses to glucoprivation, including increases in epinephrine secretion, glucocorticoid secretion, sex behavior, and food intake (reviewed by Ritter et al., 2019).

Depletion of C1 neurons attenuates the salt-induced hypertension in unanesthetized rats. Ribeiro N, Martins Sá RW, Antunes VR Brain Res 1748:147107, 2020.  doi: 10.1016/j.brainres.2020.147107

Objective: To determine if the ablation of Cl neurons mitigate the high blood pressure induced by high-salt intake. Summary: Data show that hypertension induced by high-salt intake is dependent on Cl neurons.  Dose: Bilateral injections of 2.4 ng/100 nl of Anti-DBH-SAP. The total number of TH+ neurons in the AS region was reduced by 37 ± 13% in the anti-DBH-SAP group when compared to control.  The injection of anti-Df3H-SAP into the RVLM reduced 80% of the TH+ neurons in the total number of Cl neurons, and 38% in the AS region.

Noradrenaline signaling in the LPBN mediates amylin’s and salmon calcitonin’s hypophagic effect in male rats. Boccia L, Le Foll C, Lutz TA FASEB J 34(11):15448-15461, 2020.  doi: 10.1096/fj.202001456RRR

Objective: To assess the phenotype of amylin activated LPBN (lateral parabrachial nucleus) neurons, especially to confirm the CGRPergic phenotype and to uncover the specific role of NA (noradrenaline) signaling from the AP to the LPBN. Summary: The present study confirmed the central role of the LPBN in propagating amylin’s and sCT’s hypophagic action, and particularly the importance of AP → LPBN NA signaling in the mediation of this process, through the activation of LPBN (CGRP and non-CGRP) neurons. Dose: A specific lesion of about half of the total AP NA population with the immunotoxin anti-DBH-SAP) was sufficient to abolish the hypophagic effect of amylin in rats and also reduced amylin’s ability to induce c-Fos expression in AP neurons. The neuronal pathways used to process the physiological response to amylin were investigated using 50-ng injections of Anti-DBH-SAP (Cat. #IT-03) into the area postrema (AP) or 25 ng into the lateral parabrachial nucleus (Potes et al., 2010).

Loss of diffuse noxious inhibitory control after traumatic brain injury in rats: A chronic issue. Irvine KA, Sahbaie P, Ferguson AR, Clark JD Exp Neurol 333:113428, 2020.  doi: 10.1016/j.expneurol.2020.113428

Summary: The authors hypothesize that dysfunctional descending noradrenergic and serotonergic pain control circuits may contribute to the loss of diffuse noxious inhibitory control (DNIC), a critical endogenous pain control mechanism, weeks to months after traumatic brain injury (TBI). Results suggest that TBI causes maladaptation of descending nociceptive signaling mechanisms and changes in the function of both adrenergic and serotonergic circuits. Such changes could predispose those with TBI to chronic pain. Dose: Anti-DBH-SAP (5 μg/5 μl) was injected in the left ventricle. Lesion of the LC resulted in failure of DNIC, an effect that mimics what is observed behaviorally after chronic TBI.

Adrenergic supersensitivity and impaired neural control of cardiac electrophysiology following regional cardiac sympathetic nerve loss. Tapa S, Wang L, Francis Stuart SD, Wang Z, Jiang Y, Habecker BA, Ripplinger CM Sci Rep 10:18801, 2020.  doi: 10.1038/s41598-020-75903-y

Summary: The authors present a novel mouse model of regional cardiac sympathetic hypo-innervation utilizing Anti-DBH-SAP. Dose: Either 5μL of 40 ng/μL  Anti-DBH-SAP or Mouse IgG-SAP (control) was applied three times directly to the exposed apical/anterior surface of the heart.

Neuroendocrine and Behavioral Consequences of Hyperglycemia in Cancer. Vasquez JH, Borniger JC Endocrinology 161(5):bqaa047, 2020.  doi: 10.1210/endocr/bqaa047

Summary: Ablation of norepinephrine containing projections to the arcuate (via saporin-conjugated antidopamine beta-hydroxylase injections) alters AgRP and NPY concentrations, leading to impairments in hypoglycemic (glucoprivic) or ghrelin-induced feeding. Dose: Anti-DBH-SAP (bilateral 42-ng intracranial injections) was used in rats to investigate the role of hindbrain catecholamine afferents in increased ARC NPY and AGRP gene expression (Fraley et al., 2003).

Corticolimbic stress regulatory circuits, hypothalamo–pituitary–adrenocortical adaptation, and resilience. Herman JP Chen A (Ed.): Stress Resilience 291-309, 2020.  Academic Press doi: 10.1016/B978-0-12-813983-7.00019-7

Summary: Review. Immunolesion of paraventricular nucleus (PVN)-projecting norepinephrine (NE) neurons with Anti-DBH-SAP attenuates acute stress reactivity (interestingly, to restraint), but it does not inhibit somatic or HPA axis responses to stress in any simple way (Flak et al., 2014). PVN-projecting norepinephrine (NE) neurons appear to be responsible for acute responses to systemic stressors, but they do not appear to be important in mediating effects of chronic stress (Ritter et al., 2003). Dose: Flak et al. injected 8.82 ng of Anti-DBH-SAP into the PVN.  Ritter et al. injected 42 ng into the PVN.

IT-06 IT-33 Mac-1-SAP

Inflammatory macrophages facilitate mechanical stress-induced osteogenesis. Zhang F, Huan L, Xu T, Li G, Zheng B, Zhao H, Guo Y, Shi J, Sun J, Chen A Aging (Albany NY) 12(4):3617-3625, 2020.  doi: 10.18632/aging.102833

Summary: In a mouse model of distraction osteogenesis (DO), there was significant increase in macrophages in the regeneration area. This suggests that targeting inflammatory macrophages may help to improve clinical bone repair. Dose: For saporin-mediated depletion of macrophages, DO-surgery-treated mice received i.v. injection of either Mac-1-SAP against the macrophage surface marker CD11b (20µg) once every 3 days, or Rat-IgG-SAP.

Spinal microglia‐neuron interactions in chronic pain. Ho, IHT, Chan, MTV, Wu, WKK, Liu, X J Leukoc Biol 108:1575-1592, 2020.  doi: 10.1002/JLB.3MR0520-695R

Summary: Spinal microglial activation is initiated shortly and persisted for more than 3 mo after partial sciatic nerve ligation.  Intrathecal injection of Mac1-SAP, a saporin-conjugated anti-CD11b antibody to deplete microglia, abolished cold and mechanical allodynia for 2–12 wk after injury,92 supporting the role of activated microglia for chronic pain maintenance. (Cites Echeverry S, Shi XQ, Yang M, et al. Spinal microglia are required for long-term maintenance of neuropathic pain. Pain. 2017;158(9):1792-1801.)

IT-11 SSP-SAP

Episodic stimulation of central chemoreceptor neurons elicits disordered breathing and autonomic dysfunction in volume overload heart failure. Díaz HS, Andrade DC, Toledo C, Pereyra KV, Schwarz KG, Díaz-Jara E, Lucero C, Arce-Álvarez A, Schultz HD, Silva JN, Takakura AC, Moreira TS, Marcus NJ, Del Rio R Am J Physiol Lung Cell Mol Physiol 318(1):L27-L40, 2020.  doi: 10.1152/ajplung.00007.2019

Objective: To determine the role of the central chemoreflex in the development of respiratory and autonomic dysfunction in volume overload heart failure (HF). Summary: Episodic hypercapnic stimulation triggers ventilatory plasticity and elicits cardiorespiratory  abnormalities in HF that are largely dependent on retrotrapezoid nucleus (RTN) chemoreceptor neurons. Dose: Bilateral injections of SSP-SAP, 0.6 ng/30 nL, into the RTN were administered to destroy chemoreceptor neurons.

Central circuit mechanisms of itch. Chen XJ, Sun YG Nat Commun 11:3052, 2020.  doi: 10.1038/s41467-020-16859-5

The authors summarize the progress in elucidating the neural circuit mechanism of itch at spinal and supraspinal levels.

A role for neurokinin 1 receptor expressing neurons in the paratrigeminal nucleus in bradykinin‐evoked cough in guinea‐pigs. Driessen AK, McGovern AE, Behrens R, Moe AAK, Farrell MJ, Mazzone SB J Physiol 598(11):2257-2275, 2020.  doi: 10.1113/JP279644

Objective: This study aimed to assess the involvement of paratrigeminal neurokinin 1 receptor neurons in the regulation of cough, breathing and airway defensive  responses. Summary: These findings warrant further investigations into targeting the jugular ganglia and paratrigeminal nucleus as a therapy for treating cough in disease. Dose: Targeted toxin lesions across three sites of the paratrigeminal nucleus (200nl per injection site).

Structural and Functional Consequences of Targeted Hippocampal Gaba Neuron Ablation by Stable Substance P-Saporin in Rats. Chun E Morehouse School of Medicine , 2020.

See: Chun E, Bumanglag AV, Burke SN, Sloviter RS. Targeted hippocampal GABA neuron ablation by Stable Substance P-saporin causes hippocampal sclerosis and chronic epilepsy in rats. (2019) Epilepsia 60(5):e52-e57. doi: 10.1111/epi.14723 (link)

IT-14 CTB-SAP

Adenosine 2A receptor inhibition protects phrenic motor neurons from cell death induced by protein synthesis inhibition. Seven YB, Simon AK, Sajjadi E, Zwick A, Satriotomo I, Mitchell GS Exp Neurol 323:113067, 2020.  doi: 10.1016/j.expneurol.2019.113067

Objective: To test the hypothesis that A2A receptor antagonism promotes phrenic motor neuron survival and preserves diaphragm function when faced with toxic, neurodegenerative insults that lead to phrenic motor neuron death. Summary: The authors utilized a novel neurotoxic model of respiratory motor neuron death using intrapleural injections of CTB-SAP. A2A receptors contribute to neurotoxic phrenic motor neuron death, an effect mitigated by A2A receptor antagonism. Dose: Intrapleural administration of CTB-SAP (25 μg per side) causes: 1) profound phrenic motor neuron death (~5% survival); 2) ~7-fold increase in phrenic motor neuron A2A receptor expression prior to cell death; and 3) diaphragm muscle paralysis (inactive in most rats; ~7% residual diaphragm EMG amplitude during room air breathing).

Exercise promotes recovery after motoneuron injury via hormonal mechanisms. Chew C, Sengelaub DR Neural Regen Res 15(8):1373-1376, 2020.  doi: 10.4103/1673-5374.274323

Objective: To describe how exercise is neuroprotective for motoneurons, accelerating axon regeneration following  axotomy and attenuating dendritic atrophy following the death of neighboring motoneurons. Summary: Exercise offers a simple, low barrier-to-entry behavioral intervention which is neuroprotective and pro-regenerative following neural injury. Dose: Motoneurons innervating the left vastus medialis muscle were selectively killed by intramuscular injection of CTB-SAP (2 μL, 0.1%).

Respiratory pathology in the Optn-/- mouse model of Amyotrophic Lateral Sclerosis. McCall AL, Dhindsa JS, Pucci LA, Kahn AF, Fusco AF, Biswas DD, Strickland LM, Tseng HC, ElMallah MK Respir Physiol Neurobiol 282:103525, 2020.  doi: 10.1016/j.resp.2020.103525

Summary: Tongue muscle weakness results in dysarthria and dysphagia leading to recurrent aspiration, choking, and aggravation of respiratory disease. The authors refer to work completed by Lind et al. (Lind, L.A., Murphy, E.R., Lever, T.E., Nichols, N.L., 2018. Hypoglossal Motor Neuron Death Via Intralingual CTB-saporin (CTB-SAP) Injections Mimic Aspects of Amyotrophic Lateral Sclerosis (ALS) Related to Dysphagia. Neuroscience 390, 303–316.)

Differential mechanisms are required for phrenic long-term facilitation over the course of motor neuron loss following CTB-SAP intrapleural injections. Borkowski LF, Nichols NL Exp Neurol 334:113460, 2020.  doi: 10.1016/j.expneurol.2020.113460

Objective: To understand the mechanism responsible for this difference in magnitude of phrenic long-term facilitation (pLTF) Summary: pLTF in 7d CTB-SAP treated rats is elicited primarily through TrkB and PI3K/Akt-dependent mechanisms, whereas BDNF and MEK/ERK-dependent mechanisms induce pLTF in 28d CTB-SAP treated rats. Dose: Rats received bilateral intrapleural injections of CTB-SAP; 25 μg dissolved in PBS.

Local Riluzole Release from a Thermosensitive Hydrogel Rescues Injured Motoneurons through Nerve Root Stumps in a Brachial Plexus Injury Rat Model. Fang J, Li L, Zhai H, Qin B, Quan D, Shi E, Zhu M, Yang J, Liu X, Gu L Neurochem Res 45(11):2800-2813, 2020.  doi: 10.1007/s11064-020-03120-0

The authors refer to a review by Gulino describing two rodent models of motoneuron degeneration were induced by neurotoxics including volkensin and cholera toxin-B saporin, which are able to destroy motoneurons through retrograde transportation. (Gulino R. Neuroplasticity and Repair in Rodent Neurotoxic Models of Spinal Motoneuron Disease. Neural Plast 2016:2769735. doi: 10.1155/2016/2769735)

Exercise Is Neuroprotective to Motoneuron Dendrites Following Partial Motoneuron Depletion Via a Mechanism Dependent on Androgen Receptors at the Target Muscle. Chew C Indiana Univ, Dept Psychol & Brain Sci , 2020.

See: Chew C, & Sengelaub DR. Exercise promotes recovery after motoneuron injury via hormonal mechanisms. (2020). Neural regeneration research, 15 (8):1373-1376. 2020/01/31. 10.4103/1673-5374.274323 (link)

IT-16 mu p75-SAP

Identification of Multiple Targets in the Fight against Alzheimer’s Disease. Giannoni P, Fossati S, Claeysen S, Marcello E, eds Front Aging Neurosci 12:169, 2020.  doi: 10.3389/fnagi.2020.00169

Summary: A collection of 20 articles that depict a broad representation of the most impactful advances in Alzheimer’s disease (AD) comprehension and therapeutic openings.

IT-20 Orexin-B-SAP

A common neuronal mechanism of hypertension and sleep disturbances in spontaneously hypertensive rats: Role of orexinergic neurons. Cui S-Y, Huang Y-L, Cui X-Y, Zhao H-L, Hu X, Liu Y-T, Qin Y, Kurban N, Zhang Y-H Prog Neuropsychopharmacol Biol Psychiatry 100:109902, 2020.  doi: 10.1016/j.pnpbp.2020.109902

Objective: To investigate dynamic changes in sleep patterns during the development of hypertension. Summary: Although the correlation between sleep disturbances and hypertension is very complex, common mechanisms may underlie these comorbidities. Dose: Orexin-B-SAP (HCRT2-SAP) was administered in two injections/side (100 and 200 ng/0.5 μl/injection).

IT-23 Anti-SERT-SAP

Using loss- and gain-of-function approaches to target amygdala-projecting serotonergic neurons in the dorsal raphe nucleus that enhance anxiety-related and conditioned fear behaviors. Bernabe CS, Caliman IF, Truitt WA, Molosh AI, Lowry CA, Hay-Schmidt A, Shekhar A, Johnson PL J Psychopharmacol 34(4):400-411, 2020.  doi: 10.1177/0269881119900981

Objective: To investigate the role of amygdala-projecting 5-HT neurons in the DR in innate anxiety and conditioned fear behaviors. Summary: The studies support the hypothesis that amygdala-projecting 5-HT neurons in the DR represent an anxiety and fear-on network. Dose: Each rat received two bilateral microinjections per site (100 nL each, 1 μM in ACSF) of either Anti-SERT-SAP or the control Mouse IgG-SAP.

IT-40 Bombesin-SAP

Exploration of sensory and spinal neurons expressing gastrin-releasing peptide in itch and pain related behaviors. Barry DM, Liu XT, Liu B, Liu XY, Gao F, Zeng X, Liu J, Yang Q, Wilhelm S, Yin J, Tao A, Chen ZF Nat Commun 11(1):1397, 2020.  doi: 10.1038/s41467-020-15230-y

Objective: To determine the role of GRP in sensory neurons. Summary: GRP is a neuropeptide in sensory neurons for nonhistaminergic itch, and GRP sensory neurons are dedicated to itch transmission. Dose: Bombesin-SAP (200 ng/5 μL, i.t.) was injected 2 weeks prior to optical stimulation.

More than Scratching the Surface: Recent Progress in Brain Mechanisms Underlying Itch and Scratch. Liu X, Miao XH, Liu T Neurosci Bull 36(1):85-88, 2020.  doi: 10.1007/s12264-019-00352-1

GRP receptor and AMPA receptor cooperatively regulate itch-responsive neurons in the spinal dorsal horn. Kiguchi N, Uta D, Ding H, Uchida H, Saika F, Matsuzaki S, Fukazawa Y, Abe M, Sakimura K, Ko MC, Kishioka S Neuropharmacology 170:108025, 2020.  doi: 10.1016/j.neuropharm.2020.108025

Objective: To investigate the mechanisms for the activation of itch-responsive GRPR+ neurons in the spinal dorsal horn (SDH). Summary: These findings demonstrate that GRP and glutamate cooperatively regulate GRPR+ AMPAR+ neurons in SDH, mediating itch sensation. GRP–GRPR and the glutamate–AMPAR system may play pivotal roles in the spinal transmission of itch in rodents and nonhuman primates. Dose: Bombesin-SAP and Control Blank-SAP were administered i.t. (5 μg/5 μl).

IT-42 Anti-ChAT-SAP

An acetylcholine-dopamine interaction in the nucleus accumbens and its involvement in ethanol’s dopamine-releasing effect. Loftén A, Adermark L, Ericson M, Söderpalm B Addict Biol 26(3):e12959, 2020.  doi: 10.1111/adb.12959

Summary: Basal extracellular levels of dopamine within the nucleus accumbens are not sustained by muscarinic acetylcholine, whereas accumbal Cholinergic interneurons-ACh are involved in mediating ethanol-induced dopamine release. Dose: Anti-ChAT-SAP or Rabbit IgG-SAP were infused  at a flow rate of 0.05 μl/min for 10 min giving a total volume of 0.5 μl.

IT-47 Leptin-SAP

Loss of leptin receptor-expressing cells in the hindbrain decreases forebrain leptin sensitivity. Harris RBS Am J Physiol Endocrinol Metab 318(5):E806-E816, 2020.  doi: 10.1152/ajpendo.00020.2020

Objective: This study tested whether loss of hindbrain leptin receptor signaling changed sensitivity to forebrain leptin. Summary: Loss of VMH  (ventromedial nucleus of hippocampus) leptin receptor-expressing cells prevents weight loss.  The integrated response between the hindbrain and forebrain is heavily dependent upon leptin activity in the VMH. Dose: To test forebrain leptin sensitivity Leptin-SAP and Blank-SAP rats received third ventricle injections of 0, 0.05, 0.1, 0.25 or 0.5 μg leptin.

IT-69 Nppb-SAP

Spinal GRPR and NPRA Contribute to Chronic Itch in a Murine Model of Allergic Contact Dermatitis. Liu X, Wang D, Wen Y, Zeng L, Li Y, Tao T, Zhao Z, Tao A J Invest Dermatol 140(9):1856-1866.e7, 2020.  doi: 10.1016/j.jid.2020.01.016

Intervening B-Type Natriuretic Peptide Signaling for Controlling Chronic Itch. Meng J, Chen W, Wang J Brit J Pharmacol 177(5):1025-1040, 2020.  doi: 10.1111/bph.14952

Objective: Review of recent findings used to examine the role of B-type natriuretic peptide (BNP) in itch transduction and the modulation of other pururitic proteims. Summary: Mice treated with Nppb-SAP ablated 70% of the BNP receptor-positive neurons in the spinal cord (Mishra & Hoon, 2013; Pitake, Ralph, DeBrecht & Mishra, 2018).

Molecular Genetics of Kappa Opioids in Pain and Itch Sensations. Tseng PY, Hoon MA Handb Exp Pharmacol , 2020.  doi: 10.1007/164_2020_397

Summary: The authors review the functions of the kappa opioid receptor (KOR) and its endogenous agonists dynorphins in modulating itch and pain. Nppb-SAP ablation of neurons expressing the Natriuretic olypeptide B receptor greatly reduced itch responses evoked by histamine or by intrathecal administration of Nppb, suggesting that these neurons transmit itch signals from Nppb primary afferents. Also see: Mishra SK, Hoon MA (2013) The cells and circuitry for itch responses in mice. Science 340(6135):968-971. doi: 10.1126/science.1233765

Review

Saporin from Saponaria officinalis as a Tool for Experimental Research, Modeling, and Therapy in Neuroscience. Bolshakov AP, Stepanichev MY, Dobryakova YV, Spivak, YS, Markevich, VA Toxins (Basel) 12(9):546, 2020.  doi: 10.3390/toxins12090546

A review of studies where saporin-based conjugates were used to analyze cell mechanisms of sleep, general anesthesia, epilepsy, pain, and development of Parkinson’s and Alzheimer’s diseases.

Secondary Conjugates: ZAP conjugates use your secondary agent conjugated to Saporin and makes a targeted toxin for use in vitro or in vivo. Use your biotinylated targeting agent with Streptavidin-ZAP.

IT-04  Mab-ZAP

Cellular uptake of vitamin B12: Role and fate of TCblR/CD320, the transcobalamin receptor. Gick GG, Arora K, Sequeira JM, Nakayama Y, Lai SC, Quadros EV Exp Cell Res 396(1):112256, 2020.  doi: 10.1016/j.yexcr.2020.112256

The increased and sustained expression of TCblR in proliferating cells has been used to target toxins preferentially to cancer cells and can be potentially used for targeted delivery of other anti-cancer drugs. In 2010 the authors published a paper which evaluated the potential of using immunotoxins to eliminate cancer cells expressing TCb1R the authors performed a series of in vitro experiments using their monoclonal antibody plus Mab-ZAP in varying concentrations. The results indicated that this is a viable therapeutic model that causes minimal peripheral damage.

The EphA2 and cancer connection: potential for immune-based interventions. London M, Gallo E Mol Biol Rep 47(10):8037-8048, 2020.  doi: 10.1007/s11033-020-05767-y

Summary: The authors review the most current mAb-based therapies against EphA2 expressing cancers currently in pre-clinical and/or clinical stages. They reference Sakamoto et al. who performed in vitro testing of two different EPHA2 mAbs mixed with Mab-ZAP to discover their therapeutic potential against melanoma.

IT-27 Streptavidin-ZAP

ALPPL2 Is a Highly Specific and Targetable Tumor Cell Surface Antigen. Su Y, Zhang X, Bidlingmaier S, Behrens CR, Lee NK, Liu B Cancer Res 80(20):4552-4564, 2020.  doi: 10.1101/2020.01.07.898122

Objective: To evaluate therapeutic potential of ALPPL2 targeting. Summary: Exquisite tissue specificity and broad tumor type coverage suggest that ALPPL2 could be an excellent cell surface target for therapeutic development against mesothelioma. Dose: Biotinylated M25 IgG1 and Streptavidin-ZAP were mixed at a molar ratio of 1:1.

Pseudomonas Exotoxin A Based Toxins Targeting Epidermal Growth Factor Receptor for the Treatment of Prostate Cancer. Fischer A, Wolf I, Fuchs H, Masilamani AP, Wolf P Toxins (Basel) 12(12):753, 2020.  doi: 10.3390/toxins12120753

Refers to chimeric murine-human mAb cetuximab bound to Streptavidin-ZAP.  See: Yip WL, Weyergang A, Berg K, Tonnesen HH, Selbo PK (2007) Targeted Delivery and Enhanced Cytotoxicity of Cetuximab-Saporin by Photochemical Internalization in EGFR-Positive Cancer Cells. Mol Pharm 4(2):241-251.

Control Conjugates

Using loss- and gain-of-function approaches to target amygdala-projecting serotonergic neurons in the dorsal raphe nucleus that enhance anxiety-related and conditioned fear behaviors. Bernabe CS, Caliman IF, Truitt WA, Molosh AI, Lowry CA, Hay-Schmidt A, Shekhar A, Johnson PL J Psychopharmacol 34(4):400-411, 2020.  doi: 10.1177/0269881119900981

Objective: To investigate the role of amygdala-projecting 5-HT neurons in the DR in innate anxiety and conditioned fear behaviors. Summary: The studies support the hypothesis that amygdala-projecting 5-HT neurons in the DR represent an anxiety and fear-on network. Dose: Each rat received two bilateral microinjections per site (100 nL each, 1 μM in ACSF) of either Anti-SERT-SAP or the control Mouse IgG-SAP.

Loss of leptin receptor-expressing cells in the hindbrain decreases forebrain leptin sensitivity. Harris RBS Am J Physiol Endocrinol Metab 318(5):E806-E816, 2020.  doi: 10.1152/ajpendo.00020.2020

Objective: This study tested whether loss of hindbrain leptin receptor signaling changed sensitivity to forebrain leptin. Summary: Loss of VMH  (ventromedial nucleus of hippocampus) leptin receptor-expressing cells prevents weight loss.  The integrated response between the hindbrain and forebrain is heavily dependent upon leptin activity in the VMH. Dose: To test forebrain leptin sensitivity Leptin-SAP and Blank-SAP rats received third ventricle injections of 0, 0.05, 0.1, 0.25 or 0.5 μg leptin.

GRP receptor and AMPA receptor cooperatively regulate itch-responsive neurons in the spinal dorsal horn. Kiguchi N, Uta D, Ding H, Uchida H, Saika F, Matsuzaki S, Fukazawa Y, Abe M, Sakimura K, Ko MC, Kishioka S Neuropharmacology 170:108025, 2020.  doi: 10.1016/j.neuropharm.2020.108025

Objective: To investigate the mechanisms for the activation of itch-responsive GRPR+ neurons in the spinal dorsal horn (SDH). Summary: These findings demonstrate that GRP and glutamate cooperatively regulate GRPR+ AMPAR+ neurons in SDH, mediating itch sensation. GRP–GRPR and the glutamate–AMPAR system may play pivotal roles in the spinal transmission of itch in rodents and nonhuman primates. Dose: Bombesin-SAP and Control Blank-SAP were administered i.t. (5 μg/5 μl).

Spinal GRPR and NPRA Contribute to Chronic Itch in a Murine Model of Allergic Contact Dermatitis. Liu X, Wang D, Wen Y, Zeng L, Li Y, Tao T, Zhao Z, Tao A J Invest Dermatol 140(9):1856-1866.e7, 2020.  doi: 10.1016/j.jid.2020.01.016

An acetylcholine-dopamine interaction in the nucleus accumbens and its involvement in ethanol’s dopamine-releasing effect. Loftén A, Adermark L, Ericson M, Söderpalm B Addict Biol 26(3):e12959, 2020.  doi: 10.1111/adb.12959

Summary: Basal extracellular levels of dopamine within the nucleus accumbens are not sustained by muscarinic acetylcholine, whereas accumbal Cholinergic interneurons-ACh are involved in mediating ethanol-induced dopamine release. Dose: Anti-ChAT-SAP or Rabbit IgG-SAP were infused  at a flow rate of 0.05 μl/min for 10 min giving a total volume of 0.5 μl.

Cholinergic basal forebrain degeneration due to obstructive sleep apnoea increases Alzheimer’s pathology in mice. Qian L, Kasas L, Milne MR, Rawashdeh O, Marks N, Sharma A, Bellingham MC, Coulson EJ bioRxiv 2020.03.12.989848, 2020.  doi: 10.1101/2020.03.12.989848

Dose: bilateral injections of urotensin II-saporin (UII- SAP; 0.07μg/μL per site – unless stated otherwise; generous gift from Advanced Targeting Systems)

Adrenergic supersensitivity and impaired neural control of cardiac electrophysiology following regional cardiac sympathetic nerve loss. Tapa S, Wang L, Francis Stuart SD, Wang Z, Jiang Y, Habecker BA, Ripplinger CM Sci Rep 10:18801, 2020.  doi: 10.1038/s41598-020-75903-y

Summary: The authors present a novel mouse model of regional cardiac sympathetic hypo-innervation utilizing Anti-DBH-SAP. Dose: Either 5μL of 40 ng/μL  Anti-DBH-SAP or Mouse IgG-SAP (control) was applied three times directly to the exposed apical/anterior surface of the heart.

Inflammatory macrophages facilitate mechanical stress-induced osteogenesis. Zhang F, Huan L, Xu T, Li G, Zheng B, Zhao H, Guo Y, Shi J, Sun J, Chen A Aging (Albany NY) 12(4):3617-3625, 2020.  doi: 10.18632/aging.102833

Summary: In a mouse model of distraction osteogenesis (DO), there was significant increase in macrophages in the regeneration area. This suggests that targeting inflammatory macrophages may help to improve clinical bone repair. Dose: For saporin-mediated depletion of macrophages, DO-surgery-treated mice received i.v. injection of either Mac-1-SAP against the macrophage surface marker CD11b (20µg) once every 3 days, or Rat-IgG-SAP.

Saporin

Characterization of fever and sickness behavior regulated by cytokines during infection. Li W, Luo S, Wan C Behaviour 157:855-878, 2020.  doi: 10.1163/1568539X-bja10028

Summary: This study reviews the characterization of fever and sickness behavior regulated by cytokines during infection. Dose: IL-1β-saporin or unconjugated Saporin as control (icv or ip, 1.75 μg) eliminated IL-1R1-expressing cells in the hippocampus and indicated these neurons mediate the function of peripheral IL-1β induced hypophagia.

Intracellular delivery methods using biofunctional peptide-modified extracellular vesicles. Nakase I Extracell Vesicles Circ Nucleic Acids 1:44, 2020.  American Society for Exosomes and Microvesicles 2020 Annual Meeting doi: 10.20517/evcna.2020.10

Summary: Extracellular vesicles (exosomes, EVs) with encapsulation of biofunctional molecules (e.g., enzymes and genes) are highly expected to be next-generation therapeutic carriers because of their pharmaceutical advantages. Saporin-artificially encapsulated EVs with modification of the (sC18)2 peptides showed glycosaminoglycan-dependent cell-killing activity. Our experimental techniques and findings are considered to contribute to the development for EV-based intracellular delivery system via macropinocytosis.

Quantum dot conjugated saporin activates microglia and induces selective substantia nigra degeneration. Landrigan J, Dwyer Z, Beauchamp S, Rodriguez R, Fortin T, Hayley S NeuroToxicology 76:153-161, 2020.  doi: 10.1016/j.neuro.2019.11.007

Custom Conjugates

Superior mouse eosinophil depletion in vivo targeting transgenic Siglec-8 instead of endogenous Siglec-F: Mechanisms and pitfalls. Knuplez E, Krier-Burris R, Cao Y, Marsche G, O’Sullivan J, Bochner BS J Leukoc Biol 108:43-58, 2020.  doi: 10.1002/jlb.3hi0120-381r

Objective: To determine if targeting Siglec-8 with mouse IgG1 antibodies, rather than targeting Siglec-F with rat IgG antibodies, in mice transgenic for Siglec-8, will prove to be a more effective strategy for eliminating mouse eosinophils in vivo. 
Summary: This study is the first to describe a novel mouse strain of Siglec-8+F− eosinophils—a useful tool for studying human Siglec biology in preclinical models. 
Dose: Siglec-8+F− mouse eosinophils were pretreated with 10 µg/mL saporin-conjugated isotype controls (mouse IgG1 or rat IgG2), anti-Siglec-8 (2C4) or anti-Siglec-F (9C7) antibodies for 24 h.

Labeling Neuronal Proteins with Quantum Dots for Single-Molecule Imaging. Thal LB, Kovtun O, Rosenthal SJ Methods Mol Biol 2135:169-177, 2020.  doi: 10.1007/978-1-0716-0463-2_9

Objective: The authors describe how nanometer-sized fluorescent semiconductors called quantum dots (QD) can be used to label neuronal proteins in a single QD imaging format. 
Dose: Secondary antibody-conjugated QDs target membrane proteins pre-treated with SERT Mouse Monoclonal.

Cholinergic basal forebrain degeneration due to obstructive sleep apnoea increases Alzheimer’s pathology in mice. Qian L, Kasas L, Milne MR, Rawashdeh O, Marks N, Sharma A, Bellingham MC, Coulson EJ bioRxiv 2020.03.12.989848, 2020.  doi: 10.1101/2020.03.12.989848

Dose: bilateral injections of urotensin II-saporin (UII- SAP; 0.07µg/µL per site – unless stated otherwise; generous gift from Advanced Targeting Systems)

Selected Antibodies

AB-07 Anti-FGF

YY1-mediated overexpression of long noncoding RNA MCM3AP-AS1 accelerates angiogenesis and progression in lung cancer by targeting miR-340-5p/KPNA4 axis. Li X, Yu M, Yang C J Cell Biochem 121(3):2258-2267, 2020.  doi: 10.1002/jcb.29448

western (1:1000)

Downregulation of Linc-Rna Activator of Myogenesis Lncrna Participates in FGF2 Mediated-Proliferation of Human Periodontal Ligament Stem Cells. Wu X, Cao Z, Chen H, Ou Q, Huang X, Wang Y J Periodontol 91(3):422-427, 2020.  doi: 10.1002/JPER.19-0317

Western (1:1200)

AB-265 Anti-Tri-methyl Lysine

Protein lysine methylation contributes to modulating the response of sensitive and tolerant Arabidopsis species to cadmium stress. Serre NBC, Sarthou M, Gigarel O, Figuet S, Corso M, Choulet J, Rofidal V, Alban C, Santoni V, Bourguignon J, Verbruggen N, Ravanel S Plant Cell Environ 43(3):760-774, 2020.  doi: 10.1111/pce.13692

western

AB-N01AP Anti-NGFr (mu p75)

Early Forebrain Neurons and Scaffold Fibers in Human Embryos. Qin J, Wang M, Zhao T, Xiao X, Li X, Yang J, Yi L, Goffinet AM, Qu Y, Zhou L Cerebral Cortex 30(3):913–928, 2020.  doi: 10.1093/cercor/bhz136

immunofluorescence (1:600)

AB-N03 Anti-trkA

The impact of bone cancer on the peripheral encoding of mechanical pressure stimuli. Kucharczyk MW, Chisholm KI, Denk F, Dickenson AH, Bannister K, McMahon SB Pain 161(8):1894-1905, 2020.  doi: 10.1097/j.pain.0000000000001880

immunohistochemistry (1:400)

AB-N27AP Anti-Angiotensin IIR

Chronic treatment with apelin, losartan and their combination reduces myocardial infarct size and improves cardiac mechanical function. Abbasloo, E, Najafipour, H, Vakili, A Clin Exp Pharmacol Physiol 47:393-402, 2020.  doi: 10.1111/1440-1681.13195

Objective: To determine the effect of chronic pretreatment with apelin, losartan and their combination on ischemia-reperfusion (IR) injury in the isolated perfused rat heart and on the expression of apelin-13 receptor (APJ) and angiotensin type 1 receptor (AT1R) in the myocardium. Summary: Chronic pretreatment with apelin along with AT1R antagonist had more protective effects against IR injury. Dose: 1:10000

The regulation effect of WNT-RAS signaling in hypothalamic paraventricular nucleus on renal fibrosis. Zhou G, Li J, Zeng T, Yang P, Li A J Nephrol 33(2):289-297, 2020.  doi: 10.1007/s40620-019-00637-8

immunohistochemistry, western blot

Characterization of the Renin-Angiotensin System in Aged Cavernosal Tissue and its Role in Penile Fibrosis. Bragina ME, Costa-Fraga F, Sturny M, Ebadi B, Ruoccolo RT, Santos RAS, Fraga-Silva RA, Stergiopulos N J Sex Med 17:2129–2140, 2020.  doi: 10.1016/j.jsxm.2020.08.008

immunohistochemistry (1:800)

Activation of the renin-angiotensin system in high fructose-induced metabolic syndrome. Kim M, Do GY, Kim I Korean J Physiol Pharmacol 24(4):319-328, 2020.  doi: 10.4196/kjpp.2020.24.4.319

Objective: To investigate the hypothesis that high fructose intake induces activation of the renin-angiotensin systems (RAS), resulting in hypertension and metabolic syndrome. Summary: High fructose intake increased serum renin, Ang II, triglyceride, and cholesterol levels. High fructose intake increased the expression of angiotensinogen in the liver; angiotensin-converting enzyme in the lungs; and renin, angiotensin II type 1a receptor (AT1aR), and angiotensin II type 1b receptor (AT1bR) in the kidneys. High fructose intake induces activation of RAS, resulting in hypertension and metabolic syndrome. Dose: Immunohistochemistry (Anti-AT1R)

AB-N38 Anti-Melanopsin

nGnG Amacrine Cells and Brn3b-negative M1 ipRGCs are Specifically Labeled in the ChAT-ChR2-EYFP Mouse. Cui LJ, Chen WH, Liu AL, Han X, Jiang SX, Yuan F, Zhong YM, Yang XL, Weng SJ Invest Ophthalmol Vis Sci 61(2):14, 2020.  doi: 10.1167/iovs.61.2.14

The authors speculated that type II cells might be ipRGCs. This was later verified by the strong immunostaining of type II cells in response to the melanopsin antibody UF006 (100%, 141 of 141 cells collected from 5 retinas, Figs. 6B1–B3), which probes multiple ipRGC subtypes. Immunostaining 1:10000

Examination of Zinc in the Circadian System. Moshirpour M, Nakashima AS, Sehn N, Smith VM, Thackray SE, Dyck RH, Antle MC Neuroscience 432:15-29, 2020.  doi: 10.1016/j.neuroscience.2020.02.016

Objective: To examine the anatomical and functional aspects of zinc in the circadian system. Summary: Neither enhancement nor chelation of free zinc at either the SCN or IGL altered circadian responses to phase-shifting light in hamsters. Dose: Retinal immunohistochemistry included a 20-min wash in 4% PFA prior to initiation of the IHC protocol.

AB-T044 Anti-Trans-hydroxyproline

Proline Hydroxylation Primes Protein Kinases for Autophosphorylation and Activation. Lee SB, Ko A, Oh YT, Shi P, D’Angelo F, Frangaj B, Koller A, Chen EI, Cardozo T, Iavarone A, Lasorella A Mol Cell 79(3):376-389.e8, 2020.  doi: 10.1016/j.molcel.2020.06.021

Objective: To investigate the formation of prolyl-hydroxylated intermediates as a novel mechanism of kinase maturation and a general mechanism of regulation of CMGC kinases in eukaryotes. Summary: Identified a highly conserved proline in the kinase domain of CMGC kinases that is hydroxylated by the PHD1 prolyl-hydroxylase, and this event precedes tyrosine autophosphorylation. Dose: Western blot

Obesity Shapes Metabolism in the Tumor Microenvironment to Suppress Anti-Tumor Immunity. Ringel AE, Drijvers JM, Baker GJ, Catozzi A, García-Cañaveras JC, Gassaway BM, Miller BC, Juneja VR, Nguyen TH, Joshi S, Yao CH, Yoon H, Sage PT, LaFleur MW, Trombley JD, Jacobson CA, Maliga Z, Gygi SP, Sorger PK, Rabinowitz JD, Sharpe AH, Haigis MC Cell 183(7):1848-1866.e26, 2020.  doi: 10.1016/j.cell.2020.11.009

Objective: To investigate how obesity shifts the metabolic landscape of the tumor microenvironment (TME) to inhibit T cell function and promote tumor growth. Summary: High-fat diet – induced obesity impairs CD8+ T cell function in the murine TME, accelerating tumor growth. Dose: Immunoprecipitation

Vitamin C decreases VEGF expression levels via hypoxia‑inducible factor‑1α dependent and independent pathways in lens epithelial cells. Zhao L, Wang J, Zhang Y, Wang L, Yu M, Wang F Mol Med Rep 22(1):436-444, 2020.  doi: 10.3892/mmr.2020.11103

Objective: To investigate the mechanisms underlying the effects of vitamin C on the expression levels of VEGF. Summary: Vitamin C inhibits the cell proliferation and the expression levels of VEGF in lens epithelial cells following a co-treatment with the HIF-1 inhibitor. Dose: Western blot (1:500)

AB-T125 Anti-NO-L-Cysteine

A DNA-based fluorescent probe maps NOS3 activity with subcellular spatial resolution. Jani MS, Zou J, Veetil AT, Krishnan Y Nat Chem Biol 16:660–666, 2020.  doi: 10.1038/s41589-020-0491-3

Objective: Usage of novel fluorescent DNA-based probe technology to map activities of endogenous NOS3 in live cells provides a platform to discover selective regulators of NOS3 in PM and TGN. Summary: Targeting of plasma membrane (PM) and trans-Golgi network (TGN) shows that NOS3 is tenfold less active at the Golgi but its activity is essential for the structural integrity of the Golgi. Dose: immunofluorescence