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PubMed1997

Substance P analogues potentiate the pressor response to microinjection of L-glutamate into laminas I and II of the cat dorsal horn.

Microinjection of a substance P analogue (1 mM; 7 or 10 nl) into laminae I and II of the L7 dorsal horn of decerebrate cats significantly potentiated (P < 0.05) the increase in arterial pressure evoked by microinjection of L-glutamate (109 mM; 7 or 10 nl) into these spinal sites. Microinjection of the substance P analogues (i.e., GR73638 and [Sar9,Met(O2)11]-substance P) which were selective NK-1 receptor agonists, had no impact on the cardioacceleration evoked by microinjection of L-glutamate (P > 0.05). In addition, microinjection of these analogues had no effect on the modest and non-significant increase in phrenic nerve discharge evoked by L-glutamate. We conclude that stimulation of NK-1 receptors in the superficial laminae of the dorsal horn potentiates the pressor responses to microinjection of L-glutamate.

Animals↗

PubMed1999

The antinociception produced by microinjection of a cholinergic agonist in the ventromedial medulla is mediated by noradrenergic neurons in the A7 catecholamine cell group.

Activation of neurons in the ventromedial medulla by electrical stimulation or by microinjection of opioid or cholinergic agonists produces antinociception that is mediated in part by spinally-projecting noradrenergic neurons. Several lines of evidence indicate that these noradrenergic neurons are located in the pontine A7 catecholamine cell group. For example, anatomical studies have demonstrated that neurons in the ventromedial medulla project to the noradrenergic neurons in the A7 catecholamine cell group that provide the major noradrenergic innervation of the spinal cord dorsal horn. In addition, electrical and chemical stimulation of A7 neurons produces antinociception that can be reduced by intrathecal injection of alpha2-adrenoceptor antagonists. The present studies provide more direct evidence that activation of neurons in the ventromedial medulla produces antinociception by activating noradrenergic neurons in the A7 cell group. Neurons in the ventromedial medulla were stimulated by microinjecting the cholinergic agonist carbachol (5 microg) into sites in the nucleus raphe magnus or the nucleus gigantocellularis pars alpha of pentobarbital anesthetized Sprague-Dawley rats. In some experiments, the local anesthetic tetracaine (10 microg) was then microinjected near the A7 cell group to inactivate the spinally-projecting noradrenergic neurons. In other experiments, cobalt chloride (100 mM) was microinjected near the A7 cell group to block synaptic activation of spinally-projecting noradrenergic neurons. Microinjection of carbachol into sites in the ventromedial medulla produced antinociception, assessed using the tail flick test, that lasted more than 60 min. However, the effects of carbachol were attenuated by microinjection of either tetracaine or cobalt into sites near the A7 cell group neurons identified by tyrosine hydroxylase-immunoreactivity. Similar injections of tetracaine or cobalt more than 500 microm from the A7 neurons did not alter the antinociceptive effect of carbachol. These results support the conclusion that the antinociception produced by activating neurons in the ventromedial medulla is mediated in part by the subsequent activation of spinally-projecting noradrenergic neurons in the A7 cell group.

Anesthetics, Local↗

PubMed1992

Subzonal sperm microinjection in cases of severe male factor infertility and repeated in vitro fertilization failure.

OBJECTIVE: To examine (1) fertilization rates obtained with subzonal sperm microinjection when different numbers of sperm are injected into the perivitelline space; (2) when subzonal sperm microinjection is combined with dilute insemination; and (3) the association of semen quality characteristics with fertilization. DESIGN: Subzonal sperm microinjection and subzonal sperm microinjection combined with dilute insemination was performed in 109 and 41 cycles on patients in two clinical trials in Melbourne, Australia, and Bologna, Italy, respectively. PATIENT PARTICIPANTS: Couples who have experienced repeated in vitro fertilization failure or in whom the husband has severe male factor infertility. PRIMARY OUTCOME MEASURES: The number of oocytes fertilized after injection of different numbers of sperm into the perivitelline space, the number of patients transferred, and pregnancy outcome. RESULTS: The injection of multiple numbers of sperm into the perivitelline space failed to improve monospermic fertilization rates but caused an increase in polyspermic fertilization. In patients with initial semen parameters exhibiting greater than 50% motility or greater than 50% normal morphology fertilization rates were improved when subzonal sperm microinjection-treated eggs were incubated in a dilute insemination medium. Six pregnancies were obtained, two of which have progressed to term. CONCLUSIONS: When applied to male factor patients, the subzonal sperm microinjection technique results in a 14% to 15% fertilization rate. However, of the 102 embryos transferred only three (2.9%) fetal heart beats were obtained.

Female↗

PubMed2003

The rostral ventromedial medulla mediates the facilitatory effect of microinjected orphanin FQ in the periaqueductal gray on spinal nociceptive transmission in rats.

Single unit extracellular recordings from spinal dorsal horn neurons were obtained with glass micropipettes in pentobarbital-anesthetized rats. A total of 50 wide dynamic range (WDR) neurons were studied in 25 rats. Microinjection of orphanin FQ (OFQ, 0.1 microg/0.1 microl) (a potent endogenous ligand of the opioid receptor-like receptor (ORL-1)) into the ipsilateral ventrolateral parts of periaqueductal gray (vlPAG) significantly increased C-response and post-discharge activity in most of the WDR neurons. Pre-microinjection of lidocaine (4%) into the nucleus raphe magnus (NRM) (0.5 microl), ipsilateral nucleus reticularis gigantocellularis (NGC) (0.6 microl), or nucleus gigantocellularis pars alpha (NGCalpha) and nucleus reticularis paragigantocellularis lateralis (NPGL) (0.5 microl) markedly reduced intra-vlPAG microinjection of OFQ-induced facilitatory effects on nociceptive responses of WDR neurons. Furthermore, if the NRM and ipsilateral NGC were simultaneously pre-microinjected with lidocaine, the intra-vlPAG microinjection of OFQ-induced facilitation on nociceptive responses of WDR neurons was eliminated. Also, a similar effect was observed when all the NRM, ipsilateral NGC, NGCalpha and NPGL were blocked with lidocaine. No significant effect on nociceptive responses of WDR neurons per se was found after blocking the NRM, ipsilateral NGC, NGCalpha/NPGL, or all the NRM, ipsilateral NGC, and NGCalpha/NPGL with lidocaine. These results indicate that (1) the facilitatory effect evoked by microinjection of OFQ into the vlPAG on nociceptive responses of WDR neurons in the spinal dorsal horn is primarily mediated by the NRM and ipsilateral NGC; (2) the NRM, ipsilateral NGC, and NGCalpha/NPGL do not mediate tonic descending inhibition of the spinal dorsal horn neurons.

Action Potentials↗

PubMed2000

Decreased locomotor activity after microinjection of dopamine D2/D3 receptor agonists and antagonists into lobule 9/10 of the cerebellum: a D3 receptor mediated effect?

The restricted localization of dopamine (DA) D3 receptors in the rat cerebellum lobule 9/10 appears to provide a method for investigating the in vivo selectivity of dopaminergic compounds for the D3 receptor subtype. Sprague-Dawley rats implanted with a cannula aimed at lobule 9/10 were microinjected with DA receptor ligands and immediately placed into activity chambers to record their spontaneous locomotor activity for short term (0 to 20 min) and delayed (20 to 40 min) effects. The DA D2/D3 receptor agonists quinelorane (0.1 to 2.5 microg) and 7-OH-DPAT (0.1 to 10 microg) decreased locomotor activity in the first 20 min post-microinjection. In contrast, the DAD1, receptor agonist 6-Br-APB (0.1 to 10 microg) did not affect locomotor activity during this time period, but markedly increased locomotion between 20 and 40 min at the highest dose tested. The DA receptor antagonists haloperidol and raclopride (1 to 10 microg) were also found to reduce locomotor activity. Furthermore, quinelorane and 7-OH-DPAT, but not haloperidol, when microinjected into lobules 1/2 or 6/7 (where no DA D3 receptors have been detected) decreased locomotor scores. These results show that both DA receptor agonists and antagonists decrease locomotor activity when microinjected into lobule 9/10 of the cerebellum. Additionally, DA receptor agonists can reduce spontaneous locomotion when microinjected outside of lobule 9/10. This would suggest that, at least for quinelorane and 7-OH-DPAT, the locomotor decreasing effects following microinjection into cerebellar lobule 9/10 may not be mediated by activity at DA D3 receptors, and that this behavioural assay is unlikely to provide a means for studying the in vivo pharmacology of the DA D3 receptor.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗

PubMed2002

Responses to microinjections of endomorphin and nociceptin into the medullary cardiovascular areas.

1. Cardiovascular effects of microinjections of nociceptin and endomorphin-2 into the following medullary areas were studied in urethane-anaesthetized rats: chemoreceptor projection site (CPS), intermediate portion of the nucleus tractus solitarius (I-NTS), caudal ventrolateral medullary depressor area (CVLM) and rostral ventrolateral medullary pressor area (RVLM). 2. Microinjections of nociceptin or endomorphin-2 (0.6 mmol/L each) into the CPS and RVLM elicited depressor and bradycardic responses, whereas similar injections into the I-NTS and CVLM elicited pressor and tachycardic responses. 3. The mechanism of cardiovascular responses to microinjections of these opioid peptides into different medullary areas involved in cardiovascular function can be postulated as follows: the direct effect of nociceptin and endomorphin-2 on neurons is usually inhibitory. Because the activation of CPS and RVLM by microinjections of L-glutamate results in pressor and tachycardic responses, inhibition of these areas by nociceptin and endomorphin-2 elicits depressor and bradycardic responses. Similarly, activation of neurons in the I-NTS and CVLM by microinjections of L-glutamate elicits depressor and bradycardic responses. Therefore, inhibition of these areas by microinjections of these opioid peptides elicits an increase in blood pressure and heart rate.

Analgesics, Opioid↗

PubMed1990

In vitro assessment of the viability of sheep zygotes after pronuclear microinjection.

Microinjected sheep zygotes were cultured in synthetic oviduct fluid medium (SOFM) for either 1 or 3 days and their subsequent developmental capacity was compared with that of microinjected zygotes cultured in vivo. Two experiments were carried out, using zygotes microinjected with one of three gene constructs containing the CysE and CysM genes from Salmonella typhimurium. In Experiment 1, microinjected zygotes were allocated to one of three treatments: (1) immediate transfer to recipient ewes (in vivo culture) followed by recollection 1 or 3 days later and subsequent transfer of viable embryos to other recipient ewes, (2) culture in SOFM (in vitro culture) for either 1 or 3 days before transfer to recipient ewes, and (3) immediate transfer to recipient ewes without subsequent interference. Recipient ewes were slaughtered on Day 14 of pregnancy and the number of elongated conceptuses determined. Although fewer zygotes failed to divide during in vitro culture than during in vivo culture, there were, overall, no significant differences between treatments in the percentage of zygotes that developed into elongated conceptuses (32.6-50.0%). In Experiment 2, microinjected zygotes were transferred immediately to recipient ewes or cultured in vitro for either 1 or 3 days before transfer. The number of fetuses per ewe on Day 50 of pregnancy and the number of lambs delivered per ewe were recorded. Neither the percentage of recipient ewes that became pregnant (overall 114/166, 68.7%) nor the percentage of zygotes that developed into lambs (overall 186/803, 23.2%) was significantly influenced by the culture treatment or by the gene construct microinjected.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

Reconstitution of the Golgi apparatus after microinjection of rat liver Golgi fragments into Xenopus oocytes.

We have studied the reconstitution of the Golgi apparatus in vivo using an heterologous membrane transplant system. Endogenous glycopeptides of rat hepatic Golgi fragments were radiolabeled in vitro with [3H]sialic acid using detergent-free conditions. The Golgi fragments consisting of dispersed vesicles and tubules with intraluminal lipoprotein-like particles were then microinjected into Xenopus oocytes and their fate studied by light (LM) and electron microscope (EM) radioautography. 3 h after microinjection, radiolabel was observed by LM radioautography over yolk platelet-free cytoplasmic regions near the injection site. EM radioautography revealed label over Golgi stacked saccules containing the hepatic marker of intraluminal lipoprotein-like particles. At 14 h after injection, LM radioautographs revealed label in the superficial cortex of the oocytes between the yolk platelets and at the oocyte surface. EM radioautography identified the labeled structures as the stacked saccules of the Golgi apparatus, the oocyte cortical granules, and the plasmalemma, indicating that a proportion of microinjected material was transferred to the surface via the secretion pathway of the oocyte. The efficiency of transport was low, however, as biochemical studies failed to show extensive secretion of radiolabel into the extracellular medium by 14 h with approximately half the microinjected radiolabeled constituents degraded. Vinblastine (50 microM) administered to oocytes led to the formation of tubulin paracrystals. Although microinjected Golgi fragments were able to effect the formation of stacked saccules in vinblastine-treated oocytes, negligible transfer of heterologous material to the oocyte surface could be detected by radioautography. The data demonstrate that dispersed fragments of the rat liver Golgi complex (i.e., unstacked vesicles and tubules) reconstitute into stacked saccules when microinjected into Xenopus cytoplasm. After the formation of stacked saccules, reconstituted Golgi fragments transport constituents into a portion of the exocytic pathway of the host cell by a microtubule-regulated process.

Animals↗

PubMed1992

Cell cycle effects of microinjected antisense oligodeoxynucleotides to p34cdc2 kinase.

In this study the effect of antisense oligomers targeted against the mRNA transcripts of p34cdc2 kinase on G1 progression into S-phase was examined. For this purpose, antisense, sense, or nonsense oligomers were introduced directly into the cytoplasm of T98G cells grown in monolayer cultures by glass-capillary microinjection. The microinjection of antisense oligomers (but not sense or nonsense oligomers) into growth-arrested cells before serum stimulation inhibited G1 progression into S-phase. This inhibition was correlated with a reduction in the steady-state levels of nuclear p34cdc2 protein. Microinjection of antisense oligomers into cells at 2 and 6 hours after serum stimulation also resulted in a marked inhibition in the ability of cells to enter S-phase. The inhibitory effect decreased when cells were microinjected at 12 hours after serum stimulation. When cells were microinjected at 18 and 24 hours after serum stimulation, only a slight inhibition was observed. As the antisense oligomers were introduced directly into the cytoplasm of cells at each of the time points examined, the observed differences in the inhibitory effects of the antisense oligomers at later times after serum stimulation cannot be explained by differences in uptake. An alternative explanation is that after a certain threshold level of nuclear p34cdc2 protein is reached in late G1 phase; no further increase is necessary, because the cells become committed to enter S-phase. In yeast, p34cdc2 appears to play an important role in the G1/S-phase transition at a control point in late G1 phase called START (reviewed by Lewin). In mammalian cells a control point that could be equivalent to START is the "restriction point" which is defined as the time after which inhibition of protein synthesis fails to block entry into S-phase (reviewed by Pardee). The effects observed with antisense oligomers to p34cdc2 kinase are strikingly similar to what is observed when low concentrations of the drug cycloheximide are added to these cells at different times after serum stimulation; entry into S-phase is significantly inhibited when cycloheximide is added up to 12 hours postimulation. Thus, the results reported in this study are in agreement with the idea that p34cdc2 kinase plays a role in the G1/S phase transition in mammalian cells. Finally, introduction of antisense oligomers directly into the cytoplasm of cells grown in monolayer cultures by glass-capillary microinjection appears to be a viable alternative to simply adding the oligomers to the culture medium.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗

PubMed2004

NMDA receptor antagonism blocks the cardiovascular responses to microinjection of trans-ACPD into the NTS of awake rats.

The possible interaction of glutamatergic metabotropic agonists and N-methyl-d-aspartate (NMDA) receptors was investigated in the nucleus tractus solitarii (NTS) of awake rats. The cardiovascular responses to unilateral microinjection of trans-1-amino-1,3-cyclopentanediocarboxylic acid (trans-ACPD; 250 pmol/50 nL) into the NTS (n= 8) produced hypotension (-64 +/- 4 mmHg) and bradycardic (-206 +/- 11 bpm) responses, which were blocked by previous microinjection of 2-amino-5-phosphonovaleric acid (AP-5; 10 nmol/50 nL), a selective antagonist of NMDA ionotropic receptors, into the same site. Intravenous injection of methyl-atropine blocked both the bradycardic and hypotensive responses to microinjection of trans-ACPD into the NTS, indicating that the hypotension was secondary to the intense bradycardic response. The data also showed that the bradycardic and hypotensive responses to microinjection of an NMDA agonist (10 pmol/50 nL) into the NTS were not affected by previous microinjection of alpha-methyl-4-carboxyphenylglycine (MCPG; 5 nmol/50 nL), a non-selective antagonist of metabotropic receptors. The results showing that the cardiovascular responses to microinjection of trans-ACPD into the NTS were blocked by AP-5 indicate that the responses to metabotropic agonists in the NTS involves NMDA receptors.

Animals↗

PubMed2003

Microinjection of muscimol into raphe pallidus suppresses tachycardia associated with air stress in conscious rats.

Sympathetically mediated tachycardia is a characteristic feature of the physiological response to emotional or psychological stress in mammals. Activation of neurons in the region of the dorsomedial hypothalamus appears to play a key role in the integration of this response. Tachycardia evoked by chemical stimulation of the dorsomedial hypothalamus can be suppressed by microinjection of the GABA(A) receptor agonist and neuronal inhibitor muscimol into the raphe pallidus (RP). Therefore, we tested the hypothesis that neuronal excitation in the RP mediates tachycardia seen in experimental air stress in rats. Microinjection of the GABA(A) receptor antagonist bicuculline methiodide (BMI) into the RP evoked increases in heart rate. At the same sites, microinjection of muscimol (80 pmol (100 nl)(-1)) had no effect on heart rate under baseline conditions but virtually abolished air stress-induced tachycardia, while microinjection of lower doses (10 or 20 pmol) produced transient but clear suppression. Microinjection of muscimol at sites outside the RP had no effect on stress-induced tachycardia, although modest suppression was apparent after injection at two sites within 500 microm of the RP. In another series of experiments, microinjection of muscimol (80 pmol (100 nl)(-1)) into the RP failed to influence the changes in heart rate produced by baroreceptor loading or unloading. These findings indicate that activity of neurons in the RP plays a previously unrecognized role in the generation of stress-induced tachycardia.

Air↗

PubMed1993

Opposing gastric motor responses to TRH and substance P on their microinjection into nucleus raphe obscurus of rats.

Little is known about the functional role of putative neurotransmitters in the nucleus raphe obscurus (NRO) in the control of gastric motor function, although thyrotropin-releasing hormone (TRH) and substance P (SP) have been detected in the cell bodies and/or fibers of this nucleus. Therefore, we investigated the effects of microinjection of these peptides (in a volume of 60 nl) into the caudal NRO of alpha-chloralose-anesthetized rats while recording intragastric pressure, pyloric and greater curvature motility, and blood pressure. L-Glutamate (30 nmol) was first microinjected into the NRO to identify the "gastric" region of the NRO and elicited significant increases in intragastric pressure as well as pyloric and greater curvature motility in all 16 animals. TRH (2-45 pmol, n = 16) microinjected into the same sites increased intragastric pressure as well as pyloric and greater curvature motility, and these effects were abolished by bilateral cervical vagotomy and atropine (0.5-1.0 mg/kg iv) but not by spinal cord transection. Microinjection of SP (45-405 pmol, n = 15) into the same sites decreased intragastric pressure; however, the inhibitory effect of SP on pyloric and greater curvature motility did not attain statistical significance. The effect of SP on intragastric pressure was completely abolished by bilateral vagotomy but not by systemic administration of atropine (1 mg/kg) or spinal cord transection. Microinjections of 45 pmol TRH and 405 pmol SP just outside of the NRO did not result in changes in gastric function. No overall significant changes in blood pressure were noted after microinjection of L-glutamate, TRH, or SP into the gastric region of the NRO. We conclude that both TRH and SP affect gastric motor function in the caudal NRO via a vagally mediated pathway; TRH apparently activates vagal cholinergic pathways, but the mechanism of SP-evoked gastric motor inhibition remains to be further investigated.

Animals↗

PubMed2004

Depressor and bradycardic responses to microinjections of endomorphin-2 into the NTS are mediated via ionotropic glutamate receptors.

The presence of endomorphin-like immunoreactivity has been reported in the nucleus tractus solitarius (NTS). It was hypothesized that endomorphins may play a role in cardiovascular regulation in the medial subnucleus of the NTS (mNTS). Endomorphin-2 (E-2, 0.1-4 mmol/l) was microinjected (100 nl) into the mNTS of urethane-anesthetized, artificially ventilated, adult male Wistar rats. E-2 (0.2 mmol/l) elicited decreases in mean arterial pressure (40 +/- 3.5 mmHg) and heart rate (50 +/- 7.0 beats/min). These responses were blocked by prior microinjections of naloxonazine (1 mmol/l) into the mNTS. Responses to microinjections of E-2 into the mNTS were abolished by prior combined microinjections of d-2-amino-7-phosphonoheptanoic acid (an NMDA receptor antagonist, 5 mmol/l) and 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide disodium (a non-NMDA receptor antagonist, 2 mmol/l) into the mNTS. These results were confirmed by extracellular neuronal recordings. Blockade of GABA receptors in the mNTS by prior combined microinjections of gabazine (a GABA(A) receptor antagonist, 2 mmol/l) and 2-hydroxysaclofen (a GABA(B) receptor antagonist, 100 mmol/l) also blocked the responses to E-2. It was concluded that 1) the depressor and bradycardic responses to microinjections of E-2 into the mNTS are mediated via micro(1)-opioid receptors as well as ionotropic glutamate receptors, 2) GABAergic neurons in the mNTS, which may inhibit the release of glutamate from nerve terminals, are inhibited by E-2 via micro(1)-opioid receptors, and 3) disinhibition caused by the inhibition of GABAergic neurons by E-2 may result in an increase in the glutamate release from nerve terminals, which, in turn, may elicit depressor and bradycardic responses.

Analgesics, Opioid↗

PubMed1990

Microinjection of substance P and ACh into rat intermediolateral nucleus elicits cardiovascular responses.

The effects of substance P (SP) or acetylcholine (ACh) microinjected into the intermediolateral nucleus of the spinal cord (IMLn) on arterial pressure (AP) and heart rate (HR) were investigated in 22 male Wistar rats under urethan and artificial ventilation. L-Glutamate (Glu) was microinjected into the IMLn between C7 and T4 to locate cardiovascular sites. Micropipettes containing Glu were stereotaxically positioned in 82 histologically verified sites in the IMLn between C7 and T4 on both the right and left sides. Microinjection of 4-10 nl of 0.18 M Glu in 30 of 39 explored sites at the T2 level elicited significant increases in HR (+24.2 +/- 3.1 beats/min). These changes were accompanied by significant increases in mean AP (+11.4 +/- 1.2 mmHg) at the T2 level (32/47 sites). Microinjection of 4-10 nl of SP (3 X 10(-7) to 3 X 10(-4) M) or ACh (0.005-0.5 M) in the right IMLn at the T2 level elicited increases in HR but did not affect AP. The duration of the responses to SP or ACh was significantly longer than the duration of the responses to Glu. The responses to ACh could be blocked by prior microinjection of 5 X 10(-2) M atropine. No responses were ever obtained in the left IMLn by microinjection of Glu, SP, or ACh. These results support the hypothesis that Glu, ACh, and SP mediate sympathoexcitation in the IMLn of the rat and that these excitatory responses have different temporal patterns.

Acetylcholine↗

PubMed1996

Glutamate microinjections in cerebellar cortex reproduce cerebrovascular effects of parallel fiber stimulation.

Electrical stimulation of cerebellar parallel fibers releases glutamate and increases local blood flow (BFcrb), an effect in part mediated by glutamate-induced nitric oxide (NO) production. We studied whether local microinjection of glutamate into the cerebellar cortex would produce increases in BFcrb comparable to those elicited by parallel fiber stimulation. In halothane-anesthetized rats equipped with a cranial window, glutamate was microinjected into the cerebellar molecular layer, and BFcrb was monitored by laser-Doppler flowmetry. Glutamate microinjections increased BFcrb dose dependently (2-200 pmol in 200 nl) (n = 9) and by 55 +/- 6% at 200 pmol (mean +/- SE). The magnitude and temporal profile of the increases in BFcrb compared favorably with the increase in flow produced by parallel fiber stimulation. The glutamate-induced BFcrb increase was attenuated by superfusion with the Na2+ channel blocker tetrodotoxin (10 microM; -50 +/- 10%; n = 5; P < 0.05; t-test) or by blocking synaptic activity by treatment of the cerebellar cortex with Ringer containing 20 mM Mg2+ and 0 mM Ca2+ (-80 +/- 4%; n = 6; P < 0.05). The glutamate-receptor antagonist kynurenate (10 mM) attenuated the increase in BFcrb by 59 +/- 6% (P < 0.05; n = 5). The relatively selective inhibitor of neuronal NO synthase 7-nitroindazole (100 mg/kg ip) reduced the flow response evoked by microinjection of glutamate (-46 +/- 7%; n = 5; P < 0.05) but not acetylcholine (10 microM; P > 0.05; n = 6). We conclude that glutamate microinjections increase local BFcrb via activation of glutamate receptors. The glutamate-induced vasodilation is mediated, in part, by neurally derived NO. The striking similarities between the vascular responses evoked by parallel fiber stimulation and that produced by microinjection of glutamate support the hypothesis that the increase in BFcrb produced by parallel fiber stimulation is mediated by glutamate release and activation of glutamate receptors. The data also strengthen the hypothesis that glutamate and NO are important mediators in the mechanisms linking synaptic activity to BFcrb in cerebellar cortex.

Acetylcholine↗

PubMed1984

Stimulation of luteinizing hormone release after stereotaxic microinjection of neurotensin into the medial preoptic area of rats.

Neurons immunoreactive to luteinizing hormone-releasing hormone (LHRH) are localized to the medial preoptico-septal region of the rat hypothalamus, an area known to be essential for the regulation of the surge of luteinizing hormone (LH) that precedes ovulation. The effect of neurotensin (NT) on circulating LH levels following its discrete microinjection into this region was evaluated. Within 30 min after the stereotaxic microinjection of 40 ng of NT in 50 nl into the medial preoptic area (MPOA) just rostral to the organum vasculosum of the lamina terminalis in ovariectomized (OVX), anesthetized rats, plasma LH levels rose from 417 +/- 36 to 923 +/- 113 ng/ml (p less than 0.001). There was also a significant increase (p less than 0.001) in plasma LH levels following the microinjection of 50 ng of norepinephrine in 50 nl in this same site. Similarly, levels of LH increased from 48 +/- 7 to 110 +/- 19 ng/ml (p less than 0.004) following the microinjection of NT into the rostral MPOA on the morning of proestrus in anesthetized rats. In contrast, saline, substance P, leu-enkephalin and LHRH microinjected in the rostral MPOA of OVX rats had no significant effect on circulating levels of LH. The microinjection of NT 0.4 mm rostral or caudal to this area was also without effect. The presence of two binding sites with KdS of 4.2 and 29 nM and site concentrations of 0.11 and 0.68 pmol/mg in tissue homogenates of the rostral MPOA were also established. These data suggest a role for NT in the central regulation of LH release.

Animals↗

PubMed1988

Human sperm chromosome complements after microinjection of hamster eggs.

A technique was developed for microinjection of human spermatozoa into golden hamster (Mesocricetus auratus) eggs to obtain human pronuclear chromosome complements. Before microinjection the spermatozoa were treated by brief sonication or incubation in TEST-yolk buffer to reduce motility. Very few sperm chromosome complements developed after sperm treatment with sonication and the frequency of spermatozoa with structural chromosomal abnormalities was exceedingly high (91%). The majority of sperm chromosome complements analysed had multiple breaks and rearrangements. Sperm incubation in TEST-yolk buffer before microinjection provided more analysable sperm karyotypes with a significantly lower frequency of structural chromosomal abnormalities (39%, P less than 0.001). Our results therefore suggest that sonication induces structural chromosomal abnormalities in spermatozoa. Since the frequency of chromosomal abnormalities after microinjection was higher than after sperm fertilization of hamster eggs, it appears that microinjection per se may also increase the frequency of chromosomal abnormalities in spermatozoa. These results are based on small numbers and must be confirmed on larger sample sizes, but our study suggests that microinjection of spermatozoa into eggs should not be recommended for clinical use until fully evaluated.

Animals↗

PubMed2005

Maternal aggression in Wistar rats: effect of 5-HT2A/2C receptor agonist and antagonist microinjected into the dorsal periaqueductal gray matter and medial septum.

The objective of the present study was to assess the role of the 5-HT2A/2C receptor at two specific brain sites, i.e., the dorsal periaqueductal gray matter (DPAG) and the medial septal (MS) area, in maternal aggressive behavior after the microinjection of either a 5-HT2A/2C receptor agonist or antagonist. Female Wistar rats were microinjected on the 7th postpartum day with the selective agonist alpha-methyl-5-hydroxytryptamine maleate (5-HT2A/2C) or the antagonist 5-HT2A/2C, ketanserin. The agonist was injected into the DPAG at 0.2 (N = 9), 0.5 (N = 10), and 1.0 microg/0.2 microl (N = 9), and the antagonist was injected at 1.0 microg/0.2 microl (N = 9). The agonist was injected into the medial septal area (MS) at 0.2 (N = 9), 0.5 (N = 7), and 1.0 microg/0.2 microl (N = 6) and the antagonist was injected at 1.0 microg/0.2 microl (N = 5). For the control, saline was injected into the DPAG (N = 7) and the MS (N = 12). Both areas are related to aggressive behavior and contain a high density of 5-HT receptors. Non-aggressive behaviors such as horizontal locomotion (walking) and social investigation and aggressive behaviors such as lateral threat (aggressive posture), attacks (frontal and lateral), and biting the intruder were analyzed when a male intruder was placed into the female resident's cage. For each brain area studied, the frequency of the behaviors was compared among the various treatments by analysis of variance. The results showed a decrease in maternal aggressive behavior (number of bites directed at the intruder) after microinjection of the agonist at 0.2 and 1.0 microg/0.2 microl (1.6 +/- 0.7 and 0.9 +/- 0.3) into the DPAG compared to the saline group (5.5 +/- 1.1). There was no dose-response relationship with the agonist. The present findings suggest that the 5-HT2A/2C receptor agonist has an inhibitory effect on maternal aggressive behavior when microinjected into the DPAG and no effect when microinjected into the MS. Ketanserin (1.0 microg/0.2 microl) decreased locomotion when microinjected into the DPAG and MS, but did not affect aggressive behavior. We interpret these findings as evidence for a specific role of 5-HT2A/2C receptors in the DPAG in the inhibition of female aggressive behavior, dissociated from those on motor activity.

Aggression↗